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1. class peaks constraints function sidechain not backbone and not methyl V Bld methyl all involving methyl groups V Cid The parameters A B and C are either given by the user or calculated au tomatically as follows The function calsca is used to calculate A by assuming an average dis tance of d A for all constraints from the class backbone By default the scalar B is set to B Ald in order to intersect the backbone cali bration curve at din and C is set to B 3 see also Mumenthaler et al 1997 Optionally the resulting distance constraints may be given the relative weight w Also optionally a logarithmic plot of volumes versus corre sponding minimal distances in the selected structures can be created Kd _ dnin d2 dmax 2 45 5 weight w 1 0 plot file log minimal none Derives upper distance limits from all selected peaks using a monotoni cally decreasing calibration function f d where d represents the dis tance and f d the corresponding volume e g 1 d 6 The minimal and maximal upper limit before possible pseudo atom corrections are applied are given by dnin and dmax If additional values dz are given then these discrete values are used for upper limits otherwise a contin uous calibration curve is used Optionally the resulting distance con straints may be given the relative weight w Also optionally a linear or logarithmic plot of volumes versus correspon
2. JO Uds WD 20 705 93 4 21 703 94 5 22 TIS 915 23 119 985 24 720 101 5 25 708 100 4 33 Tutorial 34 In every fourth cycle i e cycles 4 8 12 16 20 and 24 NOAH uses only the unambiguous and the ambiguous assignment lists but not the test as signment list which contains most errors Accordingly the target func tion value is usually much lower in these cycles see above while the RMSD of the resulting structure bundle may be higher for more infor mation on the internal peak lists of NOAH see command filter and Mu menthaler amp Braun 1995 In the example above 708 peaks were assigned in the H O peak list 100 of these peaks were not assigned in the reference peak list and are thus new and only 4 of 708 peaks were differently assigned in the refer ence peak list The noah grf file does also contain the corresponding numbers for the D O peak list not shown here cc Another important value is found in the files reliability x where x is again the number of the peak list These files contain the reliability of every individual assignment as well as a statistic on all peaks which are incompatible with the final structures i e where no possible assignment within the given tolerance range is compatible has a distance lt 5 A with at least one structure General reliability of structures Peaks with no ass possibility because of chemical shift 91 Unassigned
3. x y x y 0 250000 All substitutions in the command line proceed from right to left This al lows for example to compose a variable name from the values of other variables before it is used in a substitution command list_param User defined command list_param do i 1 nparam print Parameter i p i p i inserts the value of the i th command line parameter end do end list_param 17 second last Call list_param Parameter 1 17 Parameter 2 second Parameter 3 last Special variables The following variables have a special meaning for the command inter preter determines which commands are echoed i e copied to standard output before execution The possible settings are NULL off on large full OFF or not set at all In macros all commands except those built into the command line interpreter are echoed inter active commands are not echoed Commands are not echoed Both in macros and interactively all commands except those built into the command line interpreter are echoed Same as On except that the echo is surrounded by blank lines All commands are echoed and the corresponding line numbers in macros are given Same as Off except that this setting can only be overrid den by another value written in capital letters erract info nparam nproc p1 p2 A INCLAN ON Same as On except that this setting can only be overrid den by another value written in capital letters L
4. caliba 84 file Writes a chemical shift list in the format of the BioMagResBank for de tails see http www bmrb wisc edu structures n all selected structures command command anneal parameters Calculates a group of structures using the given command with optional parameters for each individual conformer If the number of structures n is specified the calculation will be performed starting from n random start conformers otherwise the calculation is performed for all selected structures Structure calculations are performed in parallel if possible dmin d 2 4 dmax d nax 5 5 vmin V 0 0 bb A calculated automatically sc B Ad methyl C B 3 weight w 1 0 avedis d 3 4 peaklist filename all peaks plot file Calibrates a peak list i e derives upper limit distance constraints from all assigned peaks and adds them to the list of current distance con straints The values dnin and dy ax give the minimal and the maximal val ue in A for a distance constraints before possible pseudo atom corrections are added Optionally only peaks with volume larger than V min Or from a peak list with given filename without extension may be considered Peaks are classified into three calibration classes peaks constraints function backbone all HN H HN H and HN H HP V Ald between residues i j with li jl lt 5 calibrate cashifts A Commands
5. noo 00000000000 00O0Oo0Oo HH H H ooooooooowoooHhHoo sooo H hb 00000000 rowuoooo ooo OOOHrONDOWONOHOOOW BR 0000000ooo0o e Rep JO BRR ooooJ 00000 0000000020 DD Ms File Formats 4 HN HAMI 0 4226 0 9063 0 000 3 0 0 0 0 5CA CAL 4530 0 0000 0 0000 3 6 7 16 0 6 HA HAL 7966 0 4689 0 9220 5 0 0 0 0 7 CB CAL 9258 0 8241 1 1992 5 8 10 12 0 8 HB HAL 5187 1 8345 1 1619 7 0 0 0 0 9 QG2 PSEUD 5244 0 0064 2 8527 0 0 0 0 0 10 061 O HYD 3 3475 0 7692 1 1194 7 11 0 0 0 11 HG1 H OXY 3 7526 1 2870 1 8728 10 0 0 0 0 12 CG2 CAL 6014 0 1530 2 5354 7 13 14 ds 0 13 HG21 HAL 3584 0 7795 3 3528 12 0 0 0 9 14 HG22 HAL 0 5230 0 0208 2 6232 12 0 0 0 9 15 HG23 H_AL 2 0917 0 8194 2 5821 12 0 0 0 9 16C CBYL 9863 1 4340 0 0000 5 17 18 0 0 170 BYL 2106 2 3886 0 0000 16 0 0 0 0 18 N LAM 3 3070 1 5402 0 0000 16 0 0 0 0 TRP RESIDUE TRP 5 28 3 27 OMEGA 2 10 0000 2 1 3 4 0 2 PHI O 0 0 0000 1 3 5 25 0 3 CHI1 Bo 113600 Be B CT Gal 25 4 CHI2 O 0 0 0000 5 7 1 de 25 5 PSI O 0 0 0000 3 5 26 28 0 G EB 0 6824 1 1357 0 0000 2 3 0 0 0 20 BYL 0 1723 2 2550 0 0000 1 0 0 0 0 3N NM 0 0000 0 0000 0 0000 1 4 5 0 0 4 HN HAMI 0 4226 0 9063 0 0000 3 0 0 0 0 5 CA CAL 1 4530 0 0000 0 0000 3 6 7 26 0 6 HA HAL 1 7862 0 5002 0 03 5 0 0 0 0 7CB CAL 2 0013 0 8067 1 17837 5 8 9 1 0 8 HB2 HAL 106202 1 8261 ler Sa O oa 0 A 9 HB3 HAL 1 6202 0 3799
6. w 00 OONNOO w 00 oouuoo 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 DADO DOA E Aa O 000 0 GOOD y ooooo0o0Ww p O ODO O GO a Qu pe pHi OOHOHNHNW 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 5577 1719 2842 4631 0000 7833 0000 9622 FD kak ZEN 0000000 ISDN ooooooa OOO OO OO N OOOO 2 0 ooooooo Ce NDI 22 EEE SD gt OS a ES Sy arl SI oooo0o0o0 oooooo0o0 oooo00o0 oooo00o0 oooo0o00 oooooo0oo0o0 00000000 Ooo Ooo a DOTO OGOOGO oooo0o0 oooo0o0 ooooo0o000 oooooo000 92295 oooooo0o0 00000 00000 Oooo fo oo 6000002 oooooo0o0o0 Ds File Formats 1 LB 0 0 0 00 2 3 5 g 0 1 Q1 PSEUD 0 0000 0 0000 0 0000 0 0 0 0 0 2 02 PSEUD 1 0000 0 0000 0 0000 0 0 0 0 0 3 Q3 PSEUD 1 0000 1 0000 0 0000 0 0 0 0 0 4 Q3 PSEUD 1 0000 1 0000 10 0000 0 0 0 0 0 5 01 PSEUD 2 0000 1 0000 0 0000 0 0 0 0 0 6 Q1 PSEUD 2 0000 1 0000 10 0000 0 0 0 0 0 702 PSEUD 2 0000 0 0000 0 0000 0 0 0 0 0 8 02 PSEUD 2 0000 0 0000 10 0000 0 0 0 0 0 903 PSEUD 3 0000 0 0000 0 0000 0 0 0 0 0 LLM2 RESIDUE LLM2 1 9 3 8 1 1B 0 0 0 00 2 3 5 7 0 101 PSEUD 0 0000 0 0000 0 0000 0 0 0 0 0 2 02
7. 10 10 281 No possible proton in dimension 1 762 Lek Qe LS 1 ba unambiguous 783 4 3 7 5 gt test al The data are peak number number of assignment possibilities in the first and in the second proton dimension number of assignment possibil ities that were discarded because of structures or transposed peaks and the resulting number of assignments In 3D lists dimension 2 is always the one coupled to the hetero atom dimension 3 regardless of what di mension 2 was in the input peak list see read peaks If a reference peak list has been loaded and the peak in consideration is assigned in this reference list NOAH will indicate that the reference assignment is either still present in the remaining assignment possibilities or that it has been discarded by NOAH The following comments may be printed at the end of each line lt UNAMBIGUOUS The assignment is unambiguous based on chemical shifts alone lt unambiguous The assignment is unambiguous only because some assignment possibilities could be discarded because of incompatibil ity with the selected structures or the absence of a transposed peak e gt test al The peak has less or equal maxamb assignment pos sibilities and was therefore taken over in the test assignment list lt ELIMINATED All assignment possibilities a peak had based on chemical shifts were eliminated because of incompatibility with the selected structures or th
8. 4019 9936 1370 0000 0659 0000 1209 288 4402 3338 2250 247 2464 0063 8967 006 0464 6047 5778 6824 1723 0000 9971 0733 0704 9681 6348 4817 3487 0465 9136 oooooooooo s 00 00 00 00 00 00 00 00 00 00 00 NONNWANNND zih 2 0000 0 9209 8447 4 e AB SI W N NRO NONNNNWNHNND Ms File Formats 9133 7265 3728 0291 0428 2067 0062 9094 1447 9346 3768 6890 8923 2067 0062 9094 1447 1357 2550 0762 8551 7127 6875 1324 4917 9366 REPND ds SHH BS beber N UY 0 010 WR J00N PUED TND NOP oOo roc OF 8 SO DO OP Nee EDI RI RE ooooooWw ee DENIED 3432 6636 9107 5373 7039 1270 5799 3466 6417 w0J0O0WRA JO 22 SD TALS 2842 3259 5214 1193 2135 2707 1041 6874 0000 8952 0000 5756 5780 6635 0464 7597 0000 0183 0000 8017 1364 0548 8814 4424 2228 7874 8681 9231 1270 5799 3466 6417 0000 0000 0000 0000 0000 0000 239A 7719 2842 4631 7833 9622 29 30 30 30 22 35 13 37 11 13 33 19 18 21 23 OFPONNAMOWWREWTONADWWWDY oooooouw oooooouw N N WODTDOWODOBONDTDOOMTOOBWOO Ww N Ww N N N oSP OCOOOVrOSO rn XO ngi e Ooo OOD a Ww w 000o0n5oo0oon Hr H N pa NA ON DANG UGD OO DANG OO OOA GO Nu ore Ww Ww N N ONG ki DAH Gg DI A
9. Initial value left bottom defines a font property with the possible values regular Regular not italics italics Italics or oblique The Symbol font is only available as regular Initial value regular determines whether the user coordinate system is automatically rescaled after reading list data The possible values are off No automatic scaling x Automatic scaling of the x dimension only y Automatic scaling of the y dimension only X y oron Automatic scaling of both dimensions If autoscaling of the x dimension is on then the values of X and X plot parameters XO and X1 are reset after reading list data such that all values in the x column of the list data are in the range between X and X If autoscaling of the y dimension is on then the values of Y and Y plot parameters YO and Y1 are reset to include all values in the y columns of the list data In general the limits are extended slightly with respect to the exact minimum and maximum in order to avoid that data points lie exactly on the margin Initial value On determines whether the border of a closed figure a rectangle a circle an ellipse a polygon a closed B zier curve or certain types of marks will be drawn as a line off Border lines are not drawn on Border lines are drawn Initial value on defines the color and can have the value black white red green blue cyan magenta or yellow All text and graphics that follows has the gi
10. Real function that returns the RMS total energy change per timestep av eraged over all timesteps of the most recently executed md command Number of the n th diastereotopic atom in the current grid search frag ment In a pair of diastereotopic atoms e g HB2 HB3 only the first one will be counted For values of n larger than the number of diastereotopic pairs in the fragment the function returns 0 Real function that returns the maximal dihedral angle change in de grees per timestep averaged over all timesteps of the most recently ex ecuted md command 127 Variables and Functions dnam n drms dval n ekin ekmean ekrms element n emean erms heavyatom n iar n iacod n iangle s iatom s iaunit n 128 Character function that returns the name of dihedral angle n Real function that returns the RMS dihedral angle change in degrees per timestep averaged over all timesteps of the most recently executed md command Real function that returns the value in degrees of dihedral angle n Real function that returns the current kinetic energy in target function units Real function that returns the mean kinetic energy averaged over all timesteps of the most recently executed md command Real function that returns the standard deviation of the kinetic energy averaged over all timesteps of the most recently executed md command Ordinal number of the atom n e g 1 for hydrogen 6
11. lt 3 1A angle selection angles of current fragment tfeut f ax 0 0 conformations name none Tries to find stereospecific assignments by systematic analysis of the lo cal conformation of amolecular fragment with grid searches If there are n pairs of stereospecificly unassigned diastereotopic substituents with in the molecular fragment 2 grid searches will be performed one for each possible combination of stereospecific assignments If a connect ed angle selection is specified then it defines the molecular fragment that will be analyzed otherwise the fragment set in the preceding grid fragment command will be used The parameter tfcut has the same meaning as in the grid search command Optionally the total number of allowed conformations can be stored in a variable with the given name Grid searches are restricted to values of the torsion angles given in the standard grid memory A on input On output the allowed values of the torsion angles are again stored in grid memory A all Deletes the selected or simply all structures from n to m name name none Copies structure n to structure m The current structure has number 0 Optionally the structure can be given a new name structure insert structure list structure select structure sort structure violate Es Commands name name Inserts the current structure into the sequence of stored structures ac
12. mat parameter is absent the program uses the format given in the peak list header line DYANAFORMAT string or if no such header line is present tries to determine the format from the peak assignments if pos sible Regardless of this input order DYANA permutes these dimen sions to hHN or hHC in 3D lists so that dimension 3 is always the heteroatom dimension and dimension 2 is the proton dimension coupled to it The option reference is used to read in a peak list as reference list for NOAH Optionally only integrated peaks i e those with an integration meth od flag different from or only assigned peaks i e those that are assigned in both proton dimensions are read Optionally the peaks are appended to those already present read peaks n15 format NhH filter overlap Reads a peak list named n15 peaks The three columns for the chemical shifts and the corresponding assignments in the peak list file refer to 15N the independent proton and the proton bound to N Peaks with comment overlap are skipped 109 Commands read prot read seq read upl read xplor 110 file file tolerance Aw 00 add Reads a XEASY chemical shift list A warning message is printed if chemical shifts are present simultaneously for an atom and its corre sponding pseudo atom Optionally only chemical shifts of currently un assigned atoms are added For chemical shifts that ar
13. 0 0 20C CBYL 1 9662 als 0 0000 5 2 22 0 0 210 OBYL 1 1773 2 3850 0 0000 20 0 0 0 0 22 N LAI 3 2853 5659 0 0000 20 0 0 0 0 ILE RESIDUE ILE 7 25 3 24 OMEGA 2 10 0000 2 1 3 4 0 2 PHI O 0 0 0000 1 3 5 23 0 PCHLI 3 1 3500 3 5 7 14 22 4 CHI22 3e T3500 ero 7 AG Ar i3 5 CHI21 3 1 3500 5 14 19 22 6 CHI31 3 1 3500 7 14 19 20 2 7 PSI O 0 0 0000 3 5 23 25 0 C CBYL 0 6824 1 1357 0 0000 2 3 0 0 0 2 0 BYL 0 1723 2 2550 0 0000 a O 0 0 0 3 N LAM 0 0000 0 0000 0 0000 1 4 5 0 0 AHN HAMI 0 4226 0 9063 0 0000 3 0 0 0 0 5 CA CAL 1 4530 0 0000 0 0000 3 6 7 23 0 6 HA HAL 1 7797 0 4805 0 9222 5 0 0 0 0 7 cB CAL 1 9888 0 8392 1 1617 5 8 10 14 0 8 HB HAL 1 6625 1 8692 1 0179 7 0 0o oO 0 9 062 PSEUD 1 2708 0 2506 2 8117 0 0 0 0 0 0 CG2 CAL 1 4086 0 3635 2 4951 7 11 12 18 0 1 HG21 HAL 1 8059 0 9770 3 3037 10 0 9 2 HG22 H_AL 0 3225 0 4528 2 4713 10 0 0 0 9 3 HG23 H_AL 1 6840 0 6781 2 6602 10 0 0 0 9 4 CGl CAL 3 5188 0 8471 1 1725 7 15 16 19 0 5 HG12 HAL 3 8906 0 1742 1 2551 14 0 0 0 17 6 HG13 H_AL 3 8906 1 2463 0 2289 14 0 0 0 17 7 061 PSEUD 3 8906 0 5361 0 7420 0 0 0 0 0 8 QDl PSEUD 4 1818 1 8856 2 6101 0 0 0 0 0 9 CDI CAL 4 0546 1 6863 2 3342 14 20 21 22 0 20 HD11 HAL 5 1444 1 6749 2 3183 19 0 0 0 18 21 HD12 HAL 3 7005 2 7124 2 2348 19 0 0 0 18 22 HD13 HAL 3 7005 1 2695 3 2771 19 0 0 0 18 23C CBYL 1 9587 4440 0 00 5 24 25 0 0 240 OBYL 1 1648 2 38385 0 0000 23 0 0 0 0 25 N LAM 3 2772 5756 0 0000 23 0 0
14. 2 PHI 0 3 CHIl 4 CHI21 5 CHI22 6 PSI 0 Cc C_BYL 20 O_BYL 3N N_AM OO GON NINH eee DAS DO AI SO Do LS O Ut o GOCO OU W w e Oo 15 WENU 00130101010 0Wwd Or wm w 0000 0000 3500 3000 0000 1 La F0 0 1 D Ls 0 l 0000 0000 3500 3000 3500 0000 FIs de 30 59 0000 3478 0000 4260 7872 3217 8009 0613 8003 1376 8004 9690 5072 1396 3162 4284 3910 5208 9505 8768 wb OOD 3986 8182 4820 9656 9585 0225 3550 0646 21 22 428 39 520 aoorono Ss JW NO hb ll ll gt 2 3 1 sli 0 0 l ooooo OOO OOO M5161 Gu A ee O a ad ob OOONDH H NHH 00003730 1278 5938 4453 5422 3049 6938 s9129 7422 8585 6453 1174 4937 3854 0000 0000 0000 18 1260 2071 0000 2981 0000 8355 2393 7456 9780 3618 1326 9864 5305 7585 7525 2047 4786 0000 0000 0000 11 13 0000 0000 0000 0000 0000 9178 2049 1839 1235 6537 2238 0214 0000 0000 0000 10 12 16 0000 0000 0000 14 15 17 17 19 19 21 23 25 25 N BWOoONoFrHORPHOADAOUNWWRENOB HH HH 13 11 12 PRP oP 0 GITA I ma ad NOT LI a 16 10 11 1 3 18 H H 000000000 JOO0DOO0OOAPRSOWOO HH H H OOPONDOOOODVOSOPOWONNOO 23 17 19 al 23 24 27 Hm DAGAN OUO ANTO H ooWUWOOOOOVOVOoSOoUIOoo
15. 4 Numbers of the two atoms m 1 2 involved in the n th distance re straint Index of dihedral angle n with respect to the original order of dihedral angles Residue index of dihedral angle n Number of the first atom belonging to residue n Number of the first dihedral angle belonging to the residue n Lower j 1 or upper j 2 bound of the i th allowed interval for angle n found by grid searches Number of allowed intervals for angle n found by grid searches Number of the dihedral angle that precedes dihedral angle n in the tree structure of dihedral angles Index of the residue with external residue number n Number of the n th selected structure Number of the last atom that is affected by a change of dihedral angle n Character function that returns the current library directory The library directory name is taken from the environment variable DYANALIB when the program starts Maximal number of structures for which the dihedral angles can be stored This value depends on the size of the protein Maximal number of structures for which the Cartesian coordinates can be stored This value depends on the size of the protein 129 Variables and Functions na naco nassign nbond n ncco nconf nd ndcdis n ndco ndcres n m ndfree nlevel nlol np_ass np_corr np_inc np_new 130 Number of atoms Number of dihedral angle constraints Number of assigned peaks Number of atoms
16. 6047 5778 3304 4085 3653 2644 2972 1756 2049 6903 0221 9484 9996 0488 OO OS OS S Om NV wa gt 00 00 142 4817 7976 0167 5255 4216 4568 2157 8362 2779 5099 1963 1027 43131 4019 9936 1370 0000 UE 0000 Les 2 3 3 4 0659 209 3304 4085 3653 2644 2972 00 00 00 00 00 00 00 00 00 00 NN oo IA DONNNDNWWWWMNMN wN WNN Oo 11 8551 7127 6875 3635 0215 5916 4863 8260 8907 3584 5959 1747 6228 4293 7403 6718 6897 1808 4100 1932 0000 9441 1782 9304 6642 9228 4021 9735 5955 5845 6890 9039 2045 0117 9094 1447 H DNNDOHOOOHrOOOHrOOoOOOOoOH N HH HWNDNDH HHNNDNWNWWNDDNDDNDOHOOOHOOOHOOOOCOH N HH HWMNDND HN W PRPRRPRPP NVWOUNWEIDAWNH al a 284 270 104 6874 0000 8952 0000 5756 5780 6635 0464 1597 0000 WN PIOWPONONUNONOWER Jou N Ne UW HHNND Wo ul 0000 8017 1493 0590 8578 1462 8279 Ww ORPORPOUONOWRRATODNADWWWNHENO WDA WERE RNs G0 Ma I OP ONOAAOWWRRTODNANDWWWDY hop H w w w NNN NNN N OSOHONDOOODO JNSOPWOSOOOOOOSNSOUNODNDOOODNS JOOPBWOO N N WOOOOV VON oOPONDOOOOSOORPRWOO NNN N MJ ONOK N N Nu H GN DEN DODO DO Dp OO OH Nw O oa Ww w ooorowooo m m N Nu Re oOoUWWOOHOOONOUWOWOOOVOVOOOU OO N Ww o a w ooooooooooooooooowooowou
17. 605 OEST 0 5255 2 5916 1 1193 3 7 0 0 7 05 CAL 2216 1 4863 1 2135 6 8 9 1 0 8 H5 HAL 2 4568 1 8260 1 2707 7 0 0 0O 10 9 H5 HAL 2157 0 8907 2 1041 7 0 0 0 10 0 05 PSEUD 18362 1 3584 1 6874 0 0 0 0 0 1 047 CAL 2779 0 5959 0 0000 7 12 13 21 0 2 Ha HAL 5099 1 1747 0 8952 11 0 0 0 0 3 c3 CAL 2 1963 0 6228 0 0000 11 14 15 37 0 443 HAL 3 1027 0 4293 0 5756 13 0 0 0 0 5c2 CAL 13131 7403 0 5780 13 16 17 19 0 6 H2 HAL 14019 6718 1 6635 15 0 0 0 0 7 02 O HYD 9936 2 8894 0 0464 15 18 0 0 0 8 HO2 HOXY 2 1370 3 5157 0 7597 17 0 0 0 0 9 01 CAL 0 0000 4100 0 0000 15 20 21 22 0 20 HI HALI 0 0659 7932 1 0183 19 0 0 0 0 2104 OEST 0 0000 0 0000 0 0000 11 19 0 0 0 22 N9 NA 1 1209 19441 0 8017 19 23 35 0 0 23 C4 CARO 2 3784 2 2371 0 3436 22 24 33 0 0 24 N3 NA 2 8170 2 0867 0 9370 23 235 0 0 0 25 C2 CARO 4 1036 2 4736 1 0196 24 26 27 0 0 26 H2 HARO 4 6227 2 4240 1 9654 25 0 0 0 0 2IN NA 4 9152 2 9484 0 0482 25 28 0 0 0 28 c6 CARO 4 4155 3 0749 2114 27 29 33 0 0 29 N6 NA 5 2378 3 5526 2 1808 28 30 31 0 0 30 H61 HA 6 1905 3 8024 9570 29 0 0 0 322 31 H62 HA 4 8967 3 6581 3 1256 29 0 32 32 06 PSEUD 5 5436 3 7303 2 5413 0 0 0 0 0 33 C5 CARO 3 0716 2 6978 4177 93 28 34 0 34 N7 NA 2 2897 2 7047 2 5414 33 35 0 0 0 35 c8 CARO 1 1186 2 2402 2 1184 22 34 36 0 0 36 H8 HARO 0 2357 2 0985 2 7240 35 0 0 0 0 37 03 OEST 2 6047 0 9094 1 3466 13 38 0 0 0 38P PAL 3 5778 2 1447 1 6417 37 0 0 0 0 CYT
18. DYANA A range Ap range condition Remove constraints for distances between atoms specified by A range and atoms specified by A range Optionally only constraints that fulfill one or several conditions are removed The format of a condition is the same as for the command distances The command can only be used for distance constraints A An range 179 COFIMA retain save sort type writeaco 180 Change the name of atoms or angles A into A gt As an example the three commands rename HB2 XXX rename HB3 HB2 rename XXX HB3 exchange the names of the atoms HB2 and HB3 A range A ranges condition Retain only constraints for distances between atoms specified A range and atoms specified by A range Optionally only constraints that fulfill one or several conditions are retained The format of a condition is the same as for the command distances The command can only be used for distance constraints Write an output Cartesian coordinate distance constraint or angle con straint file with the same name and the same format as the input file from which the data was read A A2 Az Ag range Sort atoms distance constraints or angle constraints If there are no atom or angle specifications on the command line the items are sorted according to a default order Otherwise the items are sorted into the or der given by the atom or
19. assignments aa se Minimal violation Peak proton shift in in the structure NOAH ss 0 001 i bundle A assi I number ppm both Ss g 20 CYSS HB2 9 MET A 4 20 CYSS HB2 28 CYSS HN 0 62 20 HB 29 ILE HN 0 74 re than one Cee O 791 7 nn Number of assignment Comment Assignment ISANGE possibilities based on reference chemical shift alone peak list In the above example NOAH has assigned the peak to a different proton pair than the one in the manually assigned reference peak list Howev er the both assignments seem to be compatible with some of the struc tures in the structure bundle and the peak consequently received a reliability distance of 0 A An example output illustrates some different possibilities 26 MET HN 28 CYSS HN 1 0 0 92 DOE 26 MET HN 29 ILE HN 1 5 0 61 0 794 0 0 3 More than one assignment possible 32 THR HA 32 THR HN 0 0 2523 Oz 814 100 0 1 9 MET HA 32 THR HN 6 0 0 58 1 32 THR HB 32 THR HN 9 0 1 51 Oi 816 0 0 2 More than one assignment possible 34 ASP HN 32 THR HN 2 1 1 28 de Age 817 8 0 2 6 GLU HA 8 ALA HN 3 0 0 68 0 RE 824 3 4 2 Peak 794 has two assignment possibilities but NOAH and also the spec troscopist assigned it to a short range NOE Peak 814 has only 1 possible assignment based on the chemical shifts alone and this assignment is au tomatically fulfilled because it is a intraresidual fixed distance It has theref
20. atom swap atom vdw Es Commands able tolerance The number of peaks assigned to the proton is also printed atom selection all atoms list delete Defines selected atoms as stereoassigned It is sufficient to select one atom of a diastereotopic pair to define both diastereotopic partners as ste reoassigned Optionally all stereo partners may be listed or the stereo specific assignments of selected atoms may be deleted atom selection all atoms optimal Swaps diastereotopic partners in peaks distance constraints coupling constants and chemical shifts but not in the structure itself It is suffi cient to select one atom of a diastereotopic pair to swap both diaste reotopic partners Optionally diastereotopic pairs which are not already stereoassigned may be swapped optimally in order to achieve the lowest possible target function value atom selection scale s 1 0 increment Ar 0 0 hincrement Ar 0 0 Selects atoms which are included into the van der Waals check The op tional parameters scale all selected atom radii by a factor s increment them by Ar or increment only the radii of heavy atoms with directly bound hydrogen atoms by Ar hincrement The latter parameter is used in the DYANA standard minimization procedure to compensate for the exclusion of hydrogen atoms in the lower minimization levels see macro vtfmin 83 Commands bmrblist calc_all
21. dco ps In the plot against the sequence up per distance limits are classified according to their range R white intraresidual constraints R 0 light grey sequential constraints R 1 dark grey medium range R lt 5 black long range R 5 not present in this example A FrameMaker MIF version of a plot of the short and medium range upper distance limits against the sequence that is often used to identify secondary structure elements is created by readdata helix seqplot seq mif which creates a MIF file seq mif that can be imported into FrameMaker in order to add other data such as amide proton exchange rates etc A RMSD cluster analysis that can detect whether structures are clustered in groups into distinct regions of conformation space is performed by readdata helix read_all helix cor cluster cluster The result is a Postscript file with name cluster ps The plot shows a clustering tree Along the vertical axis the structures are listed ordered according to the clustering found On the horizontal axis the minimal RMSD between any two structures in the clusters combined so far is shown 2 4 6 8 10 Ds Tutorial In addition to standard plots GRAF can be used to produce general graphics for instance plots made from tabulated data Assume that we are given a table of values AN WB 0100 al Mm OCOOWMOATNAUOBWNE Hr To produce a plot of this data the table is s
22. r r 1 Ir ral A high score means that the distance constraint is supported by other constraints while a score of O means that the constraint is isolated Deletes all distance constraints Compares distance constraints For every selected distance constraint other selected constraints to the same atom pair are searched If the in formation level is full a line is written for each comparison containing the two distances and the atom names At the end a histogram is printed with the number of constraints that were found for each difference inter val This command is useful for the comparison of two differently cali brated distance constraints files the second one must be loaded with read upl file append Deletes all selected distance constraints Keeps only the selected distance constraints Lists all selected distance constraints Adds pseudo atom corrections to all selected upper limit distance con straints Pseudo atom corrections are only added to constraints that in volve pseudo atoms The correction is given by the distance between the pseudo atom and the hydrogen atoms that it represents d first atom selection _ second atom selection weight w 1 0 lol Creates a new distance constraint an upper limit unless the option lol is set of d A between all atoms matching the first atom selection and those matching the second atom selection A weight might be specified 89 Commands A distanc
23. w w000AN Y wRARJOJ37000wwu0 RE H ooooooosorowoo6o HH oOoOoDVDVOonso gt rowommmoo N PRP PRP oOoooounoa N BR ooovovsor gt rowooooo ooroooJOoOJoOo ooovvo souoo bo e h MOYNMOWODOWOYJOYNMOO N Re ooroo J oooooov0oo000 H H oooooooooo0ooO00O00HOo0oo0oo0o0 H H ooooooooooo0o0O0O0HrO000000 oooovooooo0oo0o0 HH 0000000004000 00002000 ht 0000000000000 0000002000 Ms File Formats 1 OMEGA 2 10 0000 2 1 3 4 0 2 PHI O Of 0 0000 1 3 5 20 0 3 CHI1 3 1 35007 3 5 T 11 49 4 CHI2 0 0 0 0000 5 7 114 2 19 5 PSI O 0 0 0000 3 5 20 22 0 1C GB 0 6824 1 1357 0 000 2 3 0 0 0 20 O BYL 0 1723 2 2550 0 000 1 0 0 0 0 3N AM 0 0000 0 0000 0 0000 1 4 5 0 0 4 HN HAMI 0 4226 0 9063 0 00 3 0 0 0 0 5CA CAL 1 4530 0 0000 0 0000 3 6 7 20 0 6 HA HAL 1 7658 0 4880 0 9231 5 0 0 0 0 7 cB CAL 1 9963 0 8228 1 1699 5 8 9 11 0 8 HB2 HAL 1 6134 1 8405 1 0938 7 0 0 0 10 9 HB3 HAL 1 6134 0 4071 2 1019 7 0 0 10 0 QB PSEUD 1 6134 1 1238 1 5979 0 0 0 0 0 1 CG C VIN 3 5040 0 8713 1 2388 7 12 13 0 0 2 ND1 NAMO 4 1876 1 5636 2 2231 11 14 15 0 0 3 CD2 CARO 4 4515 0 3061 0 4368 11 16 17 0 0 4 HDI HAMI 3 7713 2 0867 2 9668 12 0 0 0 0 5 CEI CARO 5 4873 1 4156 2 0126 12 16 18 0 0 6 NE2 NAMO 5 6488 0 6364 0 9048 13 15 19 0 0 7 HD2 HARO 4 2578 0 3120 0 4399 13 0 0 0 0 8 HRI HARO 6 2871 1 8416 2 6184 15 0 0 0 9 HE2 HA 6 5236 0 3132 0 5438 16 0
24. 0 0 17 0 OL 40 05 0 13 0 0 3 0 06 0 0 1 0 05 0 0 4 0 05 0 0 0 0 04 Min 0 12 0 0 7 0 14 0 0 0 0 03 O 0 4 0 13 0 0 0 0 01 Max 0 449 Y 15857 0 33 00 045 017 Oe 1 4 00 25 0 0 1 0 12 For each structure 1ts number target function value and statistical mea sures for restraint violations are given Structures can be selected or deselected using the command structure select Most commands that act on groups of structures apply only to the selected structures The function selected i can be used to check whether structure i is selected As seen in the first section of this chapter there are several macros end ing with _all that perform actions on a group of selected structures e g calc_all 5 calculates 5 structures and stores them as structures 1 5 write_all filename writes all selected structures to disk read_all cor reads all files with the extension cor and stores them as struc tures 1 2 Even though the chemical shifts of two diastereotopic protons or methyl groups e g HP and HP in Tyr can usually be distinguished it is not always possible to obtain stereospecific assignments In such cases the usual strategy consists of provisionally assigning each one of the shifts to one of the diastereotopic partners The uncertainty of the assignment is then considered by the distance modify command which corrects i e loosens the corresponding distance constraints to allow for b
25. 0 000 CA 3 23 4 738 0 000 HA 3 24 38 080 0 000 CB 3 25 3 024 0 000 HB2 3 26 2 956 0 000 HB3 3 623 109 960 0 000 ND2 3 28 7 400 0 000 HD21 3 29 6 660 0 000 HD22 3 624 121 460 0 000 N 4 31 7 940 0 000 HN 4 32 54 370 0 000 CA 4 33 5 261 0 000 HA 4 34 43 970 0 000 CB 4 35 1 732 0 000 HB2 4 36 1 454 0 000 HB3 4 37 27 300 0 000 CG 4 38 1 522 0 000 HG 4 39 0 890 0 000 OD1 4 40 0 870 0 000 OD2 4 Peak list Peak lists follow the format used by the program XEASY Bartels et al 1995 The program DYANA can handle two dimensional homonuclear and three dimensional heteronuclear peak lists A peak list file starts with a line Number of dimensions n where n is either 2 or 3 possibly 162 Ds File Formats followed by additional comment lines starting with For each peak there is a data line possibly followed by a comment line that contains the user defined comment for the given peak Each peak data line contains the following data the peak number n chemical shifts the peak color code integer the spectrum type a string not used by DYANA the peak volume the error of the peak vol ume not used by DYANA the integration method code a character an integer not used by DYANA n atom numbers that identify atoms in the corresponding chemical shift list a zero atom number indicates a miss ing assignment and possibly additional data that is not used by DYANA An example of a two dimensional peak list is N
26. 04 5 50 5 50 314 Upper HA GLU 6 HG2 3 GLU 6 4 23 Each line in the listing of distance constraint modifications treats a pair of distance constraints in case one diastereotopic pair without ste reospecific assignment is involved or a quartet of distance constraints in case two diastereotopic pairs are involved Not all two or four distance constraints need to be present in the input of course In case a distance constraint is available from one atom to the first diastereotopic substitu ent of a prochiral centre it is listed in the column below the header A1 B1 a constraint to the second diastereotopic substituent is listed below the header A1 B2 a constraint between the second diastereotopic sub stituent of one and the first diastereotopic substituent of another prochiral centre appears under the header A2 B1 etc The four columns entitled A1 B1 A1 B2 A2 B1 and A2 B2 therefore list the input dis tance constraints with presumably arbitrary stereospecific assignment These will then be replaced by the distance constraints listed in the two columns Ai Bj and QA QB the distance limits below the heading Ai Bj will apply for the individual distances involving the diastereotopic sub stituents in case one diastereotopic pair is involved there will be two such distance constraints in case of two diastereotopic pair there will be four such distance constraints the final column indicates the limits that are imposed
27. 1 822 0 014 5 HD2 LYS 57 1 697 1 689 0 019 7 HE2 LYS 57 3 121 3 114 0 016 7 QG PRO 63 2 124 2 120 0 018 4 10 shifts with spread larger than tolerance If the library dyana lib was used which contains a table with statistical distributions of chemical shifts NOAH will print a list of all shifts from the proton list which are higher or lower than the highest and lowest val ue ever observed for that particular proton hetero atom A few shifts may deviate from these values but they should be checked carefully Running NOAH Ds Tutorial If some of the NOESY peaks are assigned the consistency between peak and proton list can be checked All spreads above the value of the vari able tolerance between the peaks assigned to the same proton are list ed A large spread may indicate that a peak is assigned to the wrong proton The command peaks deviation checks the deviation between the pro ton shifts and the peak position of all assigned peaks This command is of direct interest for NOAH users as a peak that deviates by more than Ajo variable tolerance from its chemical shifts can not be assigned cor rectly by NOAH Consequently the variable tolerance should be set to the value you inted to use for the NOAH calculation dyana gt tolerance 0 01 0 01 dyana gt peak deviation Peak Dim Deviation Atom Residue 453 4 0 017 QG2 ILE 75 528 1 0 011 HN VAL 87 LEGG E 0 013 QG1 VAL 38 1939 1 0 010 HB ILE 52 2219x 31 0
28. 33 ILE CB 9 10 HA 30 ARG OB 2 72 HA 30 ARG QD 5 80 HA 33 ILE HN 3 80 5 00E 00 Dihedral angle constraints Dihedral angle constraint files contain direct constraints on individual 164 Ds File Formats dihedral angles in the form of an allowed interval 0 0 with 0 lt 0 lt 0 360 This implies that the allowed interval must not de generate to a point A data line contains the residue number the residue name the dihedral angle name the lower and upper bounds of the al lowed interval in degrees and optionally the relative weight of the con straint The default relative weight is 1 Relative weights should be positive The weight of a constraint in the target function equals the rel ative weight times the weighting factor for the corresponding type of constraints See the following example file 32 ARG PH 904 0 99 0 32 ARG PS 100 15 0 1 00E 01 32 ARG CHI 153940 123 0 33 ILE PHI 653 0 330 33 ILE PSI 85 0 15 0 33 ILE CHI 1050 300 34 GLU PH 65 0 45 0 34 GLU PS 85 0 25 0 34 GLU CH 540 125 0 In this example the second constraint has a relative weight of 0 1 all others have the default relative weight of 1 As for distance constraint files the residue number and name need not be repeated on each data line if they are missing the corresponding data of the previous data line is assumed XPLOR distance and angle const
29. 4137 23 QH1 29 ARG 8 0710 1 7963 0 3999 24 NH2 29 ARG 6 1479 0 1775 0 6305 25 HH21 29 ARG 6 2657 0 0504 1 6154 26 HH22 29 ARG 5 4935 0 8516 0 2880 27 QH2 29 ARG 5 8796 0 4510 0 9517 28 C 29 ARG 1 5863 1 1280 2 1573 29 O 29 ARG 1 1807 0 8822 3 2923 Output Cartesian coordinate files from DYANA start with three comment lines that indicate the target function value the program version used and the number of residues and atoms listed in the coordinate file re spectively Optionally the program DYANA can also output Cartesian coordinates in the format of the Protein Data Bank Bernstein et al 1977 169 File Formats gt MS 170 COFIMA The program COFIMA coordinate file manipulation is a versatile pro gram to make simple manipulations on Cartesian coordinate distance constraint and dihedral angle constraint files The program works inter actively and allows for a variety of commands Some of the operations that can be performed with COFIMA are e Conversion between different data file formats e Renaming of atoms residues and dihedral angles e Deletion of atoms distance or angle constraints e Listing of specific atoms distance or angle constraints e Measurement of distances and dihedral angles e Attaching of atoms e g hydrogens e Insertion of pseudo atoms or pseudo atom constraints e Generation of covalent connectivities e Sorting of atoms distance or angle constraints The program consists of
30. 7993 28 C BYL 1 9838 1 4350 0 0000 290 O BYL 1 2064 2 388 0 0000 30N NAMM 3 3043 1 5436 0 0000 ARG RESIDUE ARG 7 32 3 3 OMEGA 2 10 0000 2 1 3 2 PHI O 0 0 0000 30 5 3 CHI1 3 1 3500 3 5 7 4 CHI2 3 1 3500 5 7 11 5 CHI3 3 1350 7 11 15 6 CHI4 O 0 0 0000 11 15 19 7 PSI 0 0 0 0000 3 5 30 C C BYL 0 6824 1 1357 0 0000 20 OBYL 0 1723 2 2550 0 0000 3N NM 0 0000 0 0000 0 0000 4 HN HAMI 0 4226 0 9063 0 0000 5 CA CAL 1 4530 0 0000 0 0000 6 HA HAL 1 7317 0 5213 0 9158 7CB CAL 2 0038 0 7402 1 2205 8 HB2 HAL 1 6375 1 7668 1 2235 9 HB3 HAL 1 6375 0 2685 2 1323 0 B PSEUD 1 6375 1 0177 1 6779 1 CG CAL 3 5338 0 7388 1 2182 2 HG2 HAL 3 9001 0 2878 1 2152 3 HG3 HAL 3 9001 1 2106 0 3064 4 G PSEUD 3 9001 0 4614 0 7608 5 CD CAL 4 0846 1 4791 2 4387 6 HD2 HAL 3 7230 2 5073 2 4444 7 HD3 HAL 3 7230 1 0090 3 3532 8 QD PSEUD 3 7230 1 7581 2 8988 9 NE NAM 5 5643 1 4643 2 4144 140 r H CaS CO ES 0100 0000000 NN WOO 0uno o000o000o0o0ro 0007 J3JJ00 HH DDNN A N H ooooooooooooooocvooouoooHro0o0000 H Hm H H oOOoSsoooNDooo onsorowo N WSO GTI UT E OA H rO sak al GTO Ww N N N DOGEN ONENG OOO GOGON DANG NN COOOANNDOOODOCOCOOCOOCOMNWOOF BODVCOOVCOOCC0O0O N NN ooouowNo N CA Ww NNN NbN O OLLO w H p oooovvooowooocoHhHoooooo Hm HH OrRRJOJ3 gt J000w0u E p H p H U H a o oOoOoOoosoooNDooo onsokrowo HO
31. K 1995 Statistical basis for the use of CY chemical shifts in protein structure determination J Magn Reson B 109 229 233 Mazur A K Dorofeev V E amp Abagyan R A 1991 Derivation and testing of explicit equations of motion for polymers described by internal coordi nates J Comp Phys 92 261 272 Mathiowetz A M Jain A Karasawa N amp Goddard III W A 1994 Protein simulations using techniques suitable for very large systems The cell mul tipole method for nonbond interactions and the Newton Euler inverse mass operator method for internal coordinate dynamics Proteins 20 227 224 Ms References McLachlan A D 1979 Gene duplication in the structural evolution of chy motrypsin J Mol Biol 128 49 79 Momany F A McGuire R F Burgess A W amp Scheraga H A 1975 En ergy parameters in polypeptides VII Geometric parameters partial atomic charges nonbonded interactions hydrogen bond interactions and intrinsic torsional potentials for the naturally occuring amino acids J Phys Chem 79 2361 2381 Mumenthaler C amp Braun W 1995 Automated assignment of simulated and experimental NOESY spectra of proteins by feedback filtering and self cor recting distance geometry J Mol Biol 254 465 480 Mumenthaler C Giintert P Braun W amp Wiithrich K 1997 Automated combined assignment of NOESY spectra and three dimensional protein structure determination J Biomol
32. NMR 10 351 362 Nagayama K amp Wiithrich K 1981 Structural interpretation of vincinal pro ton proton coupling constants 3J ap in the basic pancreatic trypsin inhibitor measured by two dimensional J resolved NMR spectroscopy Eur J Bio chem 115 653 657 N methy G Pottle M S amp Scheraga H A 1983 Energy parameters in polypeptides 9 Updating of geometrical parameters nonbonded interac tions and hydrogen bond interactions for the naturally occuring amino ac ids J Phys Chem 87 1883 1887 Ottiger M Szyperski T Luginb hl L Ortenzi C Luporini P Bradshaw R A amp Wiithrich K 1994 The NMR solution structure of the pheromone Er 2 from the ciliated protozoan Euplotes raikovi Protein Science 3 1515 1526 Pardi A Billeter M and Wiithrich K 1984 Calibration of the angular de pendence of the amide proton C proton coupling constants 3J HNO in a globular protein Use of na for identification of helical secondary struc ture J Mol Biol 180 741 751 Press W H Flannery B P Teukolsky S A Vetterling W T 1986 Numer ical Recipes The art of scientific computing Cambridge University Press Cambridge Singh U C Weiner P K Caldwell J W amp Kollman P A 1986 AMBER 3 0 University of California at San Francisco Spera S amp Bax A 1991 Empirical correlation between protein backbone conformation and C and CP 19C nuclear magnetic resonanc
33. PSEUD 2 0000 0 0000 0 0000 0 0 0 0 0 3 03 PSEUD 2 0000 2 0000 0 0000 0 0 0 0 0 4 03 PSEUD 2 0000 2 0000 10 0000 0 0 0 0 0 5 Q1 PSEUD 4 0000 2 0000 0 0000 0 0 0 0 0 6 Q1 PSEUD 4 0000 2 0000 10 0000 0 0 0 0 0 702 PSEUD 4 0000 0 0000 0 0000 0 0 0 0 0 8 02 PSEUD 4 0000 0 0000 10 0000 0 0 0 0 0 9 03 PSEUD 6 0000 0 0000 0 0000 0 0 0 0 0 LLM5 RESIDUE LLM5 L 9 3 8 1 LB 0 0 0 00 2 3 5 7 0 101 PSEUD 0 0000 0 0000 0 0000 0 0 0 0 0 2 02 PSEUD 5 0000 0 0000 0 0000 0 0 0 0 0 3 Q3 PSEUD 5 0000 5 0000 0 0000 0 0 0 0 0 4 Q3 PSEUD 5 0000 5 0000 10 0000 0 0 0 0 0 501 PSEUD 10 0000 5 0000 0 0000 0 0 0 0 0 6 01 PSEUD 10 0000 5 0000 10 0000 0 0 0 0 0 702 PSEUD 10 0000 0 0000 0 0000 0 0 0 0 0 8 Q2 PSEUD 10 0000 0 0000 10 0000 0 0 0 0 0 9 03 PSEUD 15 0000 0 0000 0 0000 0 0 0 0 0 LPM RESIDUE LPM L 7 3 6 1 LB 0 0 0 00 2 3 5 6 0 101 PSEUD 0 0000 0 0000 0 0000 0 0 0 0 0 2 02 PSEUD 1 0000 0 0000 0 0000 0 0 0 0 0 3 Q3 PSEUD 1 0000 1 0000 0 0000 0 0 0 0 0 4 03 PSEUD 1 0000 1 0000 10 0000 0 0 0 0 0 5 PSEUD 2 0000 1 0000 0 0000 0 0 0 0 0 60 PSEUD 2 0000 0 2300 0 0000 0 0 0 0 0 7N PSEUD 3 0920 1 7505 0 0000 0 0 0 0 0 LNM RESIDUE LNM 1 7 3 6 1 1B 0 0 0 00 2 3 5 6 0 1 Q1 PSEUD 0 0000 0 0000 0 0000 0 0 0 0 0 2 02 PSEUD 1 0000 0 0000 0 0000 0 0 0 0 0 3 Q3 PSEUD 1 0000 1 0000 0 0000 0 0 0 0 0 4 Q3 PSEUD 1 0000 1 0000 10 0000 0 0 0 0 0 5 C3 PSEUD 2 0000 1 0000 0 0000 0 0 0 0 0 6 03 PSEUD 2 0000 0 4260 0 0000 0 0 0 0 0 7P PSEUD 3 3856 1 2260
34. an overview file er2 ovw an angle file er2 ang and a coordinate file er2 cor are written The angle file er2 ang contains all 20 conformers sorted by increasing tar get function value For a later analysis all 20 conformers can be loaded into the program with the command read ang er2 ang The possibility to create new commands from existing ones by combin ing them in macros is a powerful feature of INCLAN A macro is created by saving a sequence of commands into a file with the extension dya It can be invoked in the same way as existing commands simply by typ ing its name Suppose that we want to build a macro to execute the example structure calculation in the first section of this tutorial Running DYANA The macro shall be called calculate i e it is stored in a file called calcu late dya and have two parameters the file name of the input and output files and the number of structures to calculate A first implementation is readdata pl calc all p2 write cor pl all overview pl p1 and p2 denote the two command line parameters The corresponding call of this macro in order to execute the above example is calculate helix 5 A second implementation of the calculate command uses the INCLAN command syntax to declare an interface with names and possibly de fault values of the command line parameters calculate calculate a group of structures 13 Tutorial DI
35. and er2c ovw contain ing the target functions of the calculated structures as well as the constraints violations e The redundant angle constraint file er2a aco used for the REDAC cycle For a calculation with three REDAC cycles and more minimization steps one could write redac kt 0 8 0 6 0 4 0 0 0 0 50 iter 300 800 1200 For more details on the REDAC strategy please refer to G ntert amp Wiithrich 1991 INCLAN is able to distribute a calculation over different processors of a shared memory parallel computer by virtue of parallel do loops These differ from ordinary loops only by the presence of the keyword parallel nproc 8 do i 1 20 parallel end do Parallel execution of a loop is accomplished by creating through the Unix system call fork several copies of the program in its current state These copies are identical except for the value of the loop counter the variable i in the above example and run in parallel Except for one instance of the program the main process all copies terminate after the execution of the last statement in the loop body The special variable nproc defines the maximal number of processes running in parallel By default the value of nproc is 1 1 e it is neces sary to explicitly set this variable to a value larger than one in order to execute a loop in parallel There is no mechanism within the program to return data from inside a parallel loop to the program 1 e
36. and coordinate files all input files have free format i e the exact positioning of the individual en tries on a line is not important subsequent input fields are separated by one or more blanks In all input files except residue library and Cartesian coordinate files the character indicates that the rest of the line is a comment that will be skipped by the program The program DYANA uses for all types of input and output data files de fault file name extensions if no extension is specified by the user file type default extension dihedral angle constraints dihedral angles coupling constants Cartesian coordinates residue library lower limit distance constraints orientation constraints Cartesian coordinates peak list chemical shift list residue sequence upper limit distance constraints XPLOR distance and angle constraints Formats of most files are those of already existing programs DIANA 137 File Formats 138 G ntert et al 1991a HABAS G ntert et al 1989 XEASY Eccles et al 1991 Bartels et al 1995 and XPLOR Briinger 1992 DG is the coordinate file format used by DIANA and other distance geometry pro grams PDB is the format used by the Protein Data Bank Bernstein et al 1977 Some features of PDB and XPLOR format are not supported by DYANA Residue library The residue library input file declares the atom types the nomenclature the dihedral angle definitions the
37. and structure violate Initial value 0 5 Hz Cutoff value for lower limit distance constraint violations in A Only violations larger than this value are listed in the commands structure list and structure violate Initial value 0 2 A 121 Variables and Functions cut_ori cut_tflocal cut_upl cut_vdw gridtime gridpoints hb_len hb_ang level 122 Cutoff value for orientation restraint violations in Hz Only violations larger than this value are listed with the commands structure list and structure violate Initial value 0 1 Hz Cutoff value for the maximal target function value in A that a single residue is allowed to have to be included into the angle statistics of the angstat make command Initial value 0 2 A2 Cutoff value for upper limit distance constraint violations in A Only violations larger than this value are listed with the commands structure list and structure violate Initial value 0 2 A Cutoff value for van der Waals violations in A Only violations larger than this value are listed with the commands structure list and struc ture violate Initial value 0 2 A The maximal expected computation time of a grid search If a grid search is expected to take longer than this value it is aborted Initial value 60 s The maximal expected number of grid points to be checked in a grid search If this value is exceeded the grid search will not be started Initial v
38. calibrated with the DYANA standard macro caliba after the 10th cycle if the option cal ibrate is specified For more informations on how to use this method please refer to the tu torial section Parameters This is the standard annealing protocol used by NOAH The input pa rameters correspond to those of the macro anneal This protocol uses different weights than anneal and never includes protons in the van der Waals check Parameters This is the standard minimization protocol used by NOAH The input pa rameters correspond to those of the macro vtfmin This protocol uses different weights than vtfmin and never includes protons in the van der Waals check name name structures n all selected structures range residue range all residues ang cor pdb hbond vdw full Sorts the selected structures with regard to their target function value and creates an overview file name ovw for the first of these structures If 103 Commands peak abs peak create 104 the name parameter is not specified and if a variable with name name is defined then its value is used as name Pair wise RMSDs are calculated for the given residue range see com mand rmsd The RMSD calculation can be suppressed by setting range If the range parameter is not specified and if a variable with name rmsdrange is defined then its value is used as residue range Optionally output angle ang DG coor
39. command interpreter are variables that are used and possibly set by the program not exclu sively by the user with eval set etc System variables are always glo bal There are several ways to insert the value of a variable or the result value of an expression into the command line Substitutions of the form variable or variable insert the complete value of the variable without trailing blanks into the command line Substitutions with differ from those with only if the value of the variable starts and ends with single quotes i e if it is a FORTRAN 77 character constant with the delimiting single quotes are retained in the substitution with they are removed A variable name that is im mediately followed by a letter digit or underscore character must be en closed in curly braces variable x 4 6 y 2 0 sum x y t a sum Set variables print This is t x y sum Substitute values This is a sum 4 6 2 0 6 60000 s t Create a FORTRAN 77 string from a normal variable print s s s s With and without single quotes s a sum a sum print t mer a summer Substitutions of the form variable format or variable format are used to format integer or real values of variables according to a FOR TRAN 77 format A format that contains the letter P or 1 applies to in Substring List element Function call Expression A INCLAN
40. constant file Coupling constants that involve un known residues or atoms can either cause an error a warning or can be skipped Optionally the coupling constants are appended to those already present file file structure n 1 unknown erroriwarningiskip error Reads a coordinates file in DG format If there is a DYANA header in the file the target function value will be read from the header Optionally only the n th structure may be read from a multi conformer file Other wise all m structures in a multi conformer file are read and stored as structures 1 m The first structure read will also be stored in the default structure memory O The presence of unknown residues or atoms can either cause an error a warning or they can be skipped 107 Commands read lib read lol read ori read pdb 108 file file o convert file Reads a residue library Optionally a library file with atom pointers con verted from numeric to name format or vice versa may be written see chapter File Formats file file e unknown erroriwarningliskip error append Reads a lower limit distance constraints file Constraints that involve un known residues or atoms can either cause an error a warning or can be skipped Optionally the distance constraints are appended to those already present file file m unknown erroriwarningiskip error append Reads an orientation constraint file Constra
41. cording to its target function value Optionally the structure can be given aname sum average rms Lists target function value and statistics of restraint violations for all se lected structures For each type of restraints upper distance limits lower distance limits van der Waals lower distance limits torsion angle re straints coupling constants and orientational restraints the number of violations exceeding the cutoff value variables cut_upl cut_lol etc either the sum average or rms root mean square violation and the maximal violation will be given Average and rms violation cannot be calculated for van der Waals restraints structure selection all structures first n all Selects structures according to the given structure selection see chapter Selections Optionally the selection may be restricted to the first n structures that are matched by the structure selection Sorts the selected structures according to their target function value structures n 1 delete Lists violations of distance constraints and angle constraints violations that exceed in at least n of the selected structures the cutoffs given by the variables cut_upl cut lol and cut_aco Optionally all violated con straints that are listed may be deleted structures select 1 20 structure violate structures 10 delete Deletes all distance and angle constraints that are violated in at least 10 out of the selec
42. current value of the plot parameter marksize lx y1 list mode where mark stands for either dot square caro plus cross or trian gle marks the positions given by the vector expressions x yj with the corresponding symbol If no vector expressions are specified all points of the list columns are marked If the x expression is omitted i e if only a single expression Vj is given the x coordinates are taken from the x column of the list file opens and initializes an output plot file in FrameMaker MIF format If another plot file is open when the mif command is executed it is closed See section mark X1 Y1 X2 V2 X3 V3 normal mode 67 INCLAN DI polygon ps rectangle scale 68 draws a polygon with the edges x y At least three points must be specified lx y1 list mode draws polygons with the edges given by the vector expressions X yj If no vector expressions are specified polygons are drawn through the points of all list columns If the x expression is omitted i e if only a sin gle expression y4 is given the x coordinates are taken from the x col umn of the list The number of list points must be three or more file opens and initializes an output plot file in Postscript format If another plot file is open when the pS command is executed it is closed X1 Y1 2 V2 draws a rect
43. for carbon atoms Real function that returns the mean total energy averaged over all timesteps of the most recently executed md command Real function that returns the standard deviation of the total energy av eraged over all timesteps of the most recently executed md command Number ofthe heavy atom associated with hydrogen or pseudo atom n Residue index of atom n Number of the dihedral angle that is restrained by the n th dihedral angle restraint Internal number of the dihedral angle with name s The string s consists of the angle name followed by the residue number The function returns 0 if s does not identify an existing angle Internal number of the atom with name s The string s consists of the atom name followed by the residue number The function returns 0 if s does not identify an existing atom Rigid unit number of a atom n i e the number of the dihedral angle im mediately preceding atom n ibond n m iccoa m n ida n m idcoa m n idord n idr n ifira n ifird n interval i n intervals n iprev n irnum n istruct n Ida n libdir maxang maxcor Ms Variables and Functions Number of the m th atom that is covalently bound to atom n m 1 4 The function returns 0 if less than m atoms are bound to atom n Numbers of the two atoms m 1 2 involved in the n th coupling con stant restraint Number of the m th defining atom of dihedral angle n m 1
44. gt value distance greater than value r lt value residue number difference less than value r gt value residue number difference greater than value r value residue number difference equal to value Note that no spaces are allowed within a condition For example the command distances HB HN r 1 d lt 5 lists all sequential distances shorter than 5 between Band amide pro tons The command cannot be used for angle constraints end Terminate the program A range A range condition limit Extract constraints for the distances between atoms specified by Aj range and atoms specified by A range The extracted distance con straints are appended to the current list of distance constraints Option ally only constraints that fulfill one or several conditions are extracted The format of a condition is the same as for the command distances The distance limit is set according to an optional limit specification It is possible to set the distance limit to the actual distance plus an offset by using the expression l offset or to set the distance limit to the smallest possible value from a list of limits by using the expression I lt j 5 lp or to set the distance limit to the largest possible value from a list of lim its by using the expression l gt l5 In Note that no spaces are allowed within these limit expressions This command can only be used with co ordinate files Display help information Instead of h
45. lower limit distance constraints Initial value 1 0 Weight value for contributions to the target function from orientational constraints Initial value 10 0 Weight value for contributions to the target function from upper limit distance constraints Initial value 1 0 Weight value for contributions to the target function from van der Waals lower limit distance constraints Initial value 2 0 Functions In the following alphabetical list of all DYANA functions arguments are denoted by n m integer r real s string x integer or real unless types are given explicitly The result type of a function is only given explicitly if it differs from the type of the argument s and if it is not obvious Several functions give access to internal data structures used by DYANA to store information about residues atoms and dihedral angles Internal ly residues atoms and dihedral angles are numbered consecutively from 1 to nr na and nd respectively The residue n atom n and dihedral angle n refer to these internal numberings For residues this internal number is called the residue index Residues have also an external residue number which is used in the input and out put of the program and which can differ from the residue index For ex ample if a sequence starts with residue ALA 101 this first residue has acoviol n r anam n angle n Angle n atom n Atom n calscale s
46. name HN resid 5 and name HB2 0 00 0 00 4 10 assign resid 5 and name HN resid 5 and name HB1 0 00 0 00 4 10 assign resid 5 and name HN resid 5 and name HB 0 00 0 00 3 36 assign resid 6 and name HN resid 6 and name HB 0 00 0 00 3 60 assign resid 6 and name HN resid 6 and name HG 0 00 0 00 3 10 assign resid 6 and name HN resid 6 and name HD1 0 00 0 00 5 30 assign resid 6 and name HN resid 6 and name HD2 0 00 0 00 5 50 total constraints 642 set echo true end set wrnlev 5 end An atom selection must select either exactly one atom or a group of at oms that is represented in DYANA by a pseudo atom Each assign state ment can define an upper limit d d and a lower limit d d_ for the corresponding distance where d d_ and d denote the three real num bers at the end of an assign statement Upper limits with d d 2 900 A and lower limits with d d_ lt 0 001 are not considered An example of an XPLOR angle constraint file is only the part printed in bold is interpreted by DYANA set message off echo off end restraints dihedral reset assign resid 2 and name C resid 3 and name N resid 3 and name CA resid 3 and name C 1 00000 240 000 85 000 2 assign resid 3 and name N resid 3 and name CA resid 3 and name C resid 4 and name N 1 00000 5 00000 100 000 2 assign resid 3 and name C resid 4 and name N resid 4 and name CA resid 4 and name C 1 00000
47. of residue 75 This command can only be used with co ordinate files Ay ri As ra Remove a specific covalent connectivity between the atom A of residue r and the atom A of residue 75 This command can only be used with coordinate files change cofima connect constraints coordinates A COFIMA Aj A range Rirl rl rl r Change residue names or residue numbers If the last parameter is DR the residue names of the specified atoms are set to R If the last parameter is r or r the residue numbers of the specified atoms are set to r If the last parameter is r or r the residue numbers of the specified atoms are incremented or decremented by r Switch to COFIMA the part of the program for the manipulation of Car tesian coordinate files Aj bj A gt b gt Generate covalent connectivities on the basis of bond length criteria This command can only be used with coordinate files Incorrect results may occur if there are large steric overlaps The command without any parameters is equivalent to the following command connect H 0 4 C 0 85 N 0 8 O 0 7 S 1 3 P 1 2 Q 999 LP 999 0 85 Usually the command can be used with these default parameters Cova lent connectivities are generated for those atom pairs with the interatom ic distance smaller than the sum of the two bond radii The bond radius of an atom is given by b if A is the leftmost atom type on the c
48. peaks 242 Incompatible peaks 38 61 per structure Histogram of displacements needed to make all peaks compatible O 1A 5 Pos 2 2 4 2 3A 10 3 4A 2 Aas DA 2 5 6A 4 6 TA 5 7 8A 2 8 9A 3 9 10A 1 From the above example we see that 91 peaks have no assignment pos sibility at all because there is no proton chemical shift within the allowed tolerance range from the peak position Furthermore there are 38 peaks which have some assignment possibilities but all of them are incompat ible with the current structure bundle 5 of these peaks could be ex plained by a slight shift of up to 1 A between the two protons of one assignment possibility These incompatible peaks are very interesting since an optimal solution Reliability of NOAH assignments Ds Tutorial together with an ideal peak list should have no incompatible peaks In practice there will always be some incompatible peaks for one or several of the following reasons 1 Some noise and spin diffusion peaks have been picked 2 Some proton chemical shifts are missing in the proton list and the NOESY peaks originating from these protons cannot be explained 3 The structures are partly distorted and can therefore not explain sev eral NOEs Experience has shown that the structures will often be distorted in pro portion to the number of incompatible peaks with significant bias when the percentage of incompatible peaks from the whole pea
49. r r cco n coord m n derms diastereotopic n dmax Ms Variables and Functions index 1 and external number 101 Real function that returns the size of the angle restraint violation for a given value r in degrees of the torsion angle n This function returns a negative result if there exists no angle restraint for angle n and 0 if the restraint s are not violated Character function that returns the name of atom n Character function that returns a string consisting of the angle name and the residue number of dihedral angle n Character function that returns a string consisting of the angle name the residue name and the residue number of dihedral angle n Character function that returns a string consisting of the atom name and the residue number of atom n Character function that returns a string consisting of the atom name the residue name and the residue number of atom n This function is used for the automatic calibration procedure included in the caliba macro The string s contains the calibration function f d re lating the peak volume f with a corresponding distance d Using this function calscale determines a scaling factor A such that the average distance bound of all peaks calibrated with the function A f d becomes r Only peaks with volume greater that r are taken into account Coupling constant value of the n th coupling constant restraint Cartesian coordinate m 1 2 3 of atom n
50. residual dipolar couplings These are related to the orientation of the corresponding chemical bond according to Eq 7 An example file with constraints for the orientation of N HN bonds 1 ARG HN 0 68 3 ASP HN 0 14 4 PHE HN 0 81 5 CYSS HN 0 94 0 05 6 LEU HN 0 42 0 05 7 GLU HN 0 52 0 05 5 0 10 TYR HN 0 95 0 05 5 0 Each line specifies in this order the following data residue number res idue name atom name residual dipolar coupling value the tolerance for the residual dipolar coupling value default value 0 1 Hz and relative weight default value 1 0 The atom that is specified must have exactly one covalent bond 167 File Formats 168 Dihedral angles Dihedral angle files are used by DYANA to store conformations in a more compact way than by storing Cartesian coordinates The format used is 13 1X A5 4 1X A5 F9 3 corresponding to the residue number and res idue name and up to four dihedral angle names and values in degrees As it is shown in the following example output dihedral angle file from DYANA the residue number and name need not to be repeated on each data line if the line corresponds to the same residue as the previous one Structure from DYANA f 2 50927E 01 29 ARG PHI 51 817 CHI 171 357 CHI2 160 460 CHI3 87 384 CHI4 85 465 PSI 56 588 30 ARG PHI 41 708 CHI 141 979 CHI2 71 718 CHI3 88 493 CHI4 83 498 PSI 50 974 31 ARG PHI 57
51. starting from AMBER DYANA FANTOM or other reasonable residue names op_am Change from OPAL to AMBER nomenclature op_di Change from OPAL to DYANA nomenclature plimits Change upper limit distance constraints from real to pseudo atoms DYANA nomenclature pseudo Insert pseudo atoms DYANA nomenclature sort Sort atoms in amino acid residues If a command should only be applied to a certain type of data files the command word may be followed with no intervening spaces by the qualifiers cofima to apply the command only to Cartesian coordi nates difima to apply the command only to distance constraints ancoma to apply the command only to angle constraints cofima to not apply the command to Cartesian coordinates difima to not apply the command to distance constraints or lancoma to not apply the command to angle constraints The following alphabetically ordered list of commands includes all commands that can be used for coordinate distance constraint and angle constraint files 141 Ap 143 44 145 range List bond angles dihedral angles or relative dihedral angles This com mand can only be used with coordinate files If three atom names are giv 173 COFIMA ancoma attach bind break 174 en the bond angle A A gt A3 is calculated If four atom names are given the dihedral angle A A7 A3 A is calculated If five atom names are given the difference b
52. structures respectively This chapter describes the syntax of the various selections used by DYANA A residue range consists of one or several of the following elements sep arated by commas m a residue number Mn a range of residue numbers Ma from the residue with number m onwards N from the first up to the residue n Atom selections have the following general form items in square brack ets are optional and items in curly braces can occur zero or more times atom residue operator atom residue where denotes negation atom denotes an atom name possibly containing wildcards or replace exactly one or any number of charac ters respectively or the word METHYL to denote all at oms in methyl groups or the word AMIDE to denote all atoms in amide groups residue denotes a residue selection A residue selection consists of one or several of the following elements name a residue name possibly containing wildcards FIRST thefirst residue 133 Selections A Angle selection Peak selection 134 LAST the last residue first the first residue of every fragment with contiguous resi due numbers last the last residue of every fragment with contiguous resi due numbers m a residue number Mn a range of residue numbers Ma from the residue with number m onwards wll from the first up to the residue n Atom selections can be combined using the following operators atoms i
53. teger numbers all other formats to real numbers x 4 6 y 2 0 sum x y print x y sum E12 3 4 6 2 0 0 660E 01 Substitutions of the form variable n m or variable n m where n and m are positive integer expressions are used to substitute with the substring between character positions n and m of the value of a variable Substring expressions can also appear on the left hand side of assign ment statements t a sum print another t 3 5 another sum t 3 program Assignment to a substring print St a program If the value of a variable is a comma separated list variable n or variable n where n is a positive integer expression substitute with the n th element of this list s 17 28 56 This is the end do i 1 length s length returns the number of elments print Element i s i end do Element 1 17 Element 2 28 Element 3 Element 4 56 Element 5 This is the end Sfunction or function substitute with the result value of a function without parameters function parameters or function parame ters substitute with the result value of a function with parameters If there are several parameters they are separated by commas x 2 5 print log x log x log x 0 916291 Hexpressiony or fexpression substitute with the result value of an expression 43 INCLAN echo 44 x 2 5 y 10 0 print x y
54. the value of the system variable tf_beta The larger the value of the long er the functional form will be close to the limiting case for small viola tions Type 1 is the normal DIANA target function Giintert ef al 1991 and type 3 is the error tolerant target function used by NOAH Mumenthaler et al 1997 tolerance tol_transp tol_una weight_aco Ms Variables and Functions Target functions of type 1 2 and 4 have unit whereas the target func tion of type 3 is dimensionless i e target function values obtained with type 3 cannot be compared with those obtained with other types In all cases the contribution to the target function from a small violation is proportional to the square of the violation For large violations the tar get function types differ significantly type 1 is proportional to d types 2 and 3 are proportional to d and type 4 is linear in d Note that distance constraints with very small upper bound b can lead to problems when the target function of type 1 is used because they get an excessive weight over other constraints To illustrate this assume two upper limit distance constraints that are violated by the same amount b 0 1 d 2 target function contribution 398 0 A b 3 1A d 5A target function contribution 6 2 A The first constraint gives a more than 60 times larger contribution than the second In such cases it is advisable to use the target function of type 2 to wh
55. the assignment is discarded In 3D peak lists the absence of expected transposed peaks may be used to eliminate wrong assignments if both protons are attached to the same hetero atom type which must correspond to the spectrum type and if both hetero atom shifts are known Pseudo atoms like QD in Phe and Tyr which represent protons attached to different hetero atoms cannot be used for this check because the position of the transposed peak is not de termined The check for transposed peaks is only performed if both residues are at least Ar positions apart in the sequence the default value of the param eter transposed means that no check for transposed peaks is done The position of the transposed peak is calculated and the peak list is screened for peaks that are positioned within tol_transp ppm of this position The assignment possibility is discarded if no such peak is found 79 Commands 80 Optionally only peaks which are already assigned in one proton dimen sion are assigned by NOAH oneass This can be useful in 3D lists where the assignment of one dimension is known and where NOAH is asked to find the assignment of the other dimension Only peaks with less than maxamb see variables possible assign ments are taken into the test assignment list With info full one output line is written for every unassigned peak 708 2 1 2 0 ALL ELIMINATED 734 de Tee 0 ALL ELIMINATED 735 1 I 1 a UNAMBIGUOUS 736 4 5
56. the backbone atoms and for all heavy atoms Optionally a residue range for the superposition may be specified For two sets of n atoms each P Y 0 the RMSD is defined by 1 1 and 41 4 gt With RMSD min Re SO 3 5 seqplot ssbond A Commands R denotes a rotation matrix and SO 3 the rotation group If the information level is full local RMSDs and global displacements are calculated for each residue Local RMSDs for residue i are calculated for the segment of n residues i n i i n n odd file file seqplot ps Analyses the upper limit distance constraints and draws a sequence plot in FrameMaker MIF if the file extension is mif or Postscript format The first three lines below the amino acid sequence represent tor sion angle restraints for the backbone torsion angles d and y and for the side chain torsion angle x For band y a triangle pointing upwards indicates a restraint that allows the torsion angle to take the values observed in an ideal a helix 57 y 47 or 349 helix 6 60 y 30 a triangle pointing downwards indi cates compatibility with an ideal parallel or antiparallel B strand 119 y 113 or 139 y 135 respectively Schultz amp Schirmer 1979 a restraint represented by a star encloses confor mations of both a and B secondary structure types and a filled cir cle marks a restraint
57. the option full is set Replace all peak volumes by their absolute value distance d 4 0 structures n additional c13 n15 Deletes current peak lists and creates expected peaks using the structures from the selected structure memories Peaks are created if the distance between two assigned proton or pseudo atom chemical shifts is less peak delete peak deviations peak distance peak list peak scale peak select peak unassign peak unique Es Commands than d in at least n of the selected structures To calculate distances where pseudo atoms are involved a r weighted average distance is deter mined between all protons that are represented by the pseudo atom With the option additional the current peaks are not deleted and the expect ed peaks are only added if they are not already present Per default a 2D peak list is created but with the options 13 or n15 a 3D 3C or 5N correlated NOESY peak list is simulated Deletes all selected peaks Prints out a list of peaks where the deviations between the peak position and the assigned chemical shift are larger than the tolerance value given by the variable tolerance If the information level is full a histogram is written at the end with the number of deviating peaks in each dimen sion for different deviations in ppm Lists for all selected peaks the average standard deviation minimum and maximum of the corresponding distance in the selected
58. the value variable substitutes the value of the variable in the command line Substitutions proceed from left to right Single quotes that delimit FOR TRAN 77 character strings are retained The curly braces in variable or variable separate the variable name variable from immediately following text expression or expressiony substitute the result value of the FORTRAN 77 expres sion variable format uses the given FORTRAN 77 format to convert the numeric value of a variable into the string that is substituted in the com mand line If the value of the variable is a comma separated list vari able n where n is an integer expression substitutes with the n th element of this list variable m n where m and n are integer expres sions substitutes with the substring between positions m and n of the val ue of the variable These three possible uses of parentheses cannot be used simultaneously separates commands that stand on the same line Note however that commands that form blocks e g do end do if end if must always appear as the first command on a line Label denotes a label that can be used as the target of a jump in a goto statement Ac treats the character c literally and allows the use of special charac ters in normal text at the end of a line indicates that the statement continues on the following line text treats text as a sin
59. van Gunsteren W F DiNola A and Haak J R 1984 Molecular dynamics with coupling to an external bath J Chem Phys 81 3684 3690 Bernstein F C Koetzle T F Williams G J B Meyer E F Jr Brice M D Rodgers J R Kennard O Shimanouchi T Tasumi M 1977 The Protein Data Bank A computer based archival file for macromolecular structures J Mol Biol 112 535 542 Braun W amp Go N 1985 Calculation of protein conformations by proton proton distance constraints A new efficient algorithm J Mol Biol 186 611 626 Braun W Bosch C Brown L R Go N amp Wiithrich K 1981 Combined use of proton proton overhauser enhancements and a distance geometry al gorithm for determination of polypeptide conformations application to mi celle bound glucagon Biochim Biophys Acta 667 377 396 Briinger A T 1992 X PLOR version 3 1 A system for X ray crystallography and NMR Yale University Press New Haven Bystrov V F 1976 Spin spin coupling and the conformational states of pep tide systems Prog NMR Spectrosc 10 41 81 Cornell W D Cieplak P Bayly C I Gould I R Merz Jr K M Ferguson D M Dpellmeyer D C Xox T Caldwell J W amp Kollman P A 1995 A second generation force field for the simulation of proteins nucleic acids and organic molecules J Am Chem Soc 117 5179 5197 DeMarco A Llin s M and W thrich K 1978a Analysis of t
60. with the following entries is given a running number the residue name the atom name the number of chemical shifts that were available for this atom the average chemical shift the standard de viation of the chemical shift the maximal chemical shift the upper and lower 5 quartiles of the chemical shift and the minimal chemical shift The Fortran format of the data lines is 5X 2A5 15 6F8 2 A similar block of data named KARPLUS is used to define Karplus type relationships of the form 37 0 A BcosO Ccos 8 8 between vicinal scalar couplings 3J and the intervening dihedral angle 6 KARPLUS 15 i HN HA 1 90 1 40 6 40 PH ZA HN C 0 10 10 4 00 PH E HN CB 0 20 1 50 4 70 PH 4 Cc 0 30 0 80 2 00 PH gt e CB 0 10 0 60 1 50 PH 6 HA 0 80 4 40 9 00 PH ie HA 0 27 0 61 0 88 PH 8 HA HB 1 80 1 60 9 50 CHI1 9 PHE HA CG 0 70 1 00 7 10 CHIL 10 TYR HA CG 0 70 1 00 7 10 CHIL del se HA CG 0 20 1 20 10 20 CHIL 2 HB 0 10 1 20 4 40 CHIl 13 HB 0 60 2 04 7 520 ICHEL 147 HB HG 1 80 1 60 9 50 CHI2 154 HB CD 0 20 1 30 10 20 CHI2 Each Karplus curve is given on one line with the following data a run ning number a residue name that may contain wildcards the two names of the atoms that are scalar coupled the parameters A B and C of the 159 File Formats 160 Karplus curve and an optional comment the corresponding dihedral an gle The Fortran format of
61. 0 The user can choose another coordinate system by specifying four pa rameters XO YO X1 and Y1 that define the coordinate values of the two reference points in the new coordinate system By default GRAF chooses after reading a table of data points a coordinate system such that all data points lie within the rectangle defined by the reference points In INCLAN string variables can be used in a similar way as in a Unix shell dyana gt name Dyana dyana gt print My name is name My name is Dyana i performs an assignment variable substitutes the value of a vari able into the command line In addition variables with numeric values can be used in expressions in the same way as in FORTRAN or other programming languages dyana gt x 7 dyana gt y 5 x dyana gt z sqrt y 10 0 dyana gt show x y z x 7 y 35 z 5 00000 show is an INCLAN command that displays the values of variables Here a different assignment sign instead of i was used Assign ments with have the meaning Evaluate the expression on the right hand side and assign the result value to the variable Note the difference to a string assignment with i dyana gt y 5 x dyana gt show y y DAK Using INCLAN control statements Ds Tutorial Expressions formed according to the rules of FORTRAN 77 may contain integer real and complex numbers logicals and character strings With in expressi
62. 0 read ori 108 read pdb 108 read peaks 109 read prot 110 read seq 110 read upl 110 read_all 111 readdata 111 redac 111 REDAC strategy 15 reliability 112 residue types ADE 150 ALA 139 ARG 140 ARG 140 ASN 141 ASP 141 ASP 141 CYS 142 CYSS 142 CYT 151 GLN 142 GLU 143 GLU 143 GLY 144 GUA 153 HIS 144 HIST 144 HIST 144 ILE 145 LEU 145 LGLY 157 LL 156 LL2 156 LLS 156 LLM 156 LLM2 157 LLMS5 157 LN 156 LNM 157 LP 156 LPM 157 LYS 146 LYS 146 MET 147 NL 155 NLM 156 PHE 147 PL 155 PLM 156 PRO 148 RADE 151 RCYT 152 RGUA 153 SER 148 THR 148 THY 154 TRP 149 TYR 149 URA 155 VAL 150 rmsd 112 Running DYANA 9 S selections angle 134 atom 133 distance constraint 135 peak 134 residue range 133 structure 135 seqplot 97 113 several molecules 27 ssbond 113 stereoassign 114 structure clear 114 structure copy 114 structure insert 115 structure list 115 structure select 115 A Index structure sort 115 structure vio 115 substitutions basic 42 expression 43 Fortran format 42 function call 43 list element 43 substring 43 sugarbond 116 sugarring 116 T torsion angle dynamics 14 translate 116 V variables global 42 local 42 special 42 system 42 vtfmin 116 W weight_aco 125 weight_cco 126 weight_lol 126 weight_ori 126 weight_upl 126 weight_vdw 126 write aco 117 write ang 118 write ass 118 write cor 118 write lol 118 write pdb 119 write peaks 119 write prot 119
63. 0 0 LEU RESIDUE LEU NE 25 OMEGA 2 10 0000 2 1 3 4 0 2 PHI 0 0 0 0000 1 3 5 24 0 3 CHI1 3 e3500 n an GR va 23 4 CHI2 3 143800 5 7 Ad ds 23 5 CHI31 3 41 3500 7 ar 15 16 18 6 CHI32 3 1 3500 7 11 19 20 22 7 PSI 0 0 0 0000 3 5 24 26 0 C CBYL 0 6824 1 1357 0 0000 2 3 0 0 0 26 BYL 0 1723 2 2550 0 0000 1 0 0 0 0 3N NM 0 0000 0 0000 0 0000 1 4 5 0 0 4 HN HAMI 0 4226 0 9063 0 000 3 0 0 0 0 5 CA CAL 1 4530 0 0000 0 0000 3 6 7 24 0 145 File Formats LYS LYS 146 D1 D2 D1 D11 D12 HD13 CD2 HD21 HD22 HD23 QOD 24 C 25 0 26 N EQOOUO II 159 000 J0O0UH4SwWN ROO JO N r NN wN RESIDUE OMEGA PHI CHI1 CHI2 CHI3 90 30014 UNO J00 PWNDHOSNINAUPwWMNH 20 HE2 21 HE3 22 QE 23 NZ 24 HZ1 25 HZ2 26 OZ 216 28 0 29 N RESIDUE OMEGA PHI CHI1 CHI2 CHI3 CHI4 CHIS PSI OPWNRP OND na U U NU pel EII ToD py ps gt ps dd cala gt a a a NA e o Na e naak a O0 ou Zoat un Powe EE LYS 0 PEREC ZEY RE Q los bs Is ys os E A o O EAE A oO I y mn un D E U os bos b Bh Gi lp oO U S ah m EEE U fe oO zeg Zoattd un Begs KK C Fed LYS 0 WRERAWWURWWUW PB as DY RAR RIE WREFPAMTADMUOUNOWWWBWWWWRPRPRPRPRERPODOOCOWWWWWON Oo ooowwWwWwWwON o aa ee 7797 9888 5941 5941 5941 29118 8975 0177 2850 9206 0080 5225 5225 1366 2196
64. 0 0 0000 3 6 8 12 0 6 HA HALI 1 7849 0 4925 0 9140 5 0 0 0 0 139 File Formats E 0850 0 9858 1 4878 9637 0 7966 1 2023 0537 0 7963 1 2019 2 1 3 10 HB2 HAL 1 6006 1 8224 1 1392 11 HB3 HAL 1 6006 0 3386 2 1223 12C C_BYL 1 9587 1 4440 0 0000 130 O BYL 1 1648 2 3835 0 0000 IAN NM 3 2772 1 5756 0 0000 ARG RESIDUE ARG 7 30 3 29 OMEGA 2 10 0000 2 1 3 2 PHI 0 0 0 0000 1 3 5 3 CHI1 3 1 3500 3 5 7 4 CHI2 3 1 3500 5 7 11 5 CHI3 3 1350 7 11 15 6 CHI4 0 0 0 0000 11 15 19 7 PSI 0 0 0 0000 3 5 28 C C BYL 0 6824 1 1357 0 0000 20 OBYL 0 1723 2 2550 0 0000 3N NM 0 0000 0 0000 0 0000 4 HN HAMI 0 4226 0 9063 0 0000 5 CA CAL 1 4530 0 0000 0 0000 6 HA HAL 1 7317 0 5213 0 9158 7 CB CAL 2 0038 0 7402 1 2205 8 HB2 HAL 1 6375 1 7668 1 2235 9 HB3 HAL 1 6375 0 2685 2 1323 0 B PSEUD 1 6375 1 0177 1 6779 1 CG CAL 3 5338 0 7388 1 2182 2 HG2 HAL 3 9001 0 2878 1 2152 3 HG3 HAL 3 9001 1 2106 0 3064 4 G PSEUD 3 9001 0 4614 0 7608 5 CD CAL 4 0846 1 4791 2 4387 6 HD2 HAL 3 7230 2 5073 2 4444 7 HD3 HAL 3 7230 1 0090 3 3532 8 QD PSEUD 3 7230 1 7581 2 8988 9 NE NAM 5 5643 1 4643 2 4144 20 HE HAM 6 0160 21 0017 1 6515 21 CZ C VIN 6 3425 2 0364 3 3576 22 NHI NAM 7 6548 1 9625 3 2357 23 HHI HAM 8 3002 2 3586 3 8888 24 NH2 N_AM 5 7870 2 6745 4 4096 25 HH21 HAM 6 3705 3 0956 5 1040 26 HH22 HAM 4 7919 2 7260 4 4945 27 QH2 PSEUD 5 5812 2 9108 4
65. 0 0000 0 0 0 0 0 LGLY RESIDUE LGLY 3 11 3 10 1 OMEGA 1 2 10 0000 2 1 3 4 0 2 PHI 0 0 0 0000 1 3 5 9 0 3 PSI 0 0 0 0000 3 5 9 11 0 alate PSEUD 0 6824 1 1357 0 0000 0 0 0 0 0 20 PSEUD 0 1723 2 2550 0 0000 0 0 0 0 0 3N PSEUD 0 0000 0 0000 0 0000 0 0 0 0 0 401 PSEUD 0 4226 0 9063 0 0000 0 0 0 0 0 5 02 PSEUD 1 4530 0 0000 0 0000 0 0 0 0 0 6 021 PSEUD 1 8202 0 5343 0 8762 0 0 0 0 0 7 022 PSEUD 1 8202 0 5343 0 8762 0 0 0 0 0 8 03 PSEUD 1 8202 0 5343 0 0000 0 0 0 0 0 IE PSEUD 2 0013 1 4284 0 0000 0 0 0 0 0 10 O PSEUD 1 2356 2 3910 0 0000 0 0 0 0 0 11 N PSEUD 3 3231 1 5208 0 0000 0 0 0 0 0 157 File Formats 158 For amino acid residues the first three characters of the name correspond to the standard three letter code the fourth character indicates positive or negative charges e g ARG or differentiates between cysteine CYS and cystine CYSS residues Cystines are involved in disulfide bridges ADE CYT GUA and THY denote the standard deoxyribonucleotides of DNA RADE RCYT RGUA and URA denote the standard nucle otides of RNA Linker residues used to treat more than one molecule and containing only pseudo atoms are also included PL to link an amino acid residue to a generic linker NL to link a nucleotide residue to a generic linker LL a generic linker residue with 1 A bond lengths and bond angles LL2 and LL5 similar linkers residue but with 2 A and 5 A bond lengths LP to link a generic linker to a following
66. 0 0000 0 0000 C HN CA 4 HN H_AM 0 4226 0 9063 0 0000 N o CA C_AL 4530 0 0000 0 0000 N HA CB C 6 HA H_AL 7416 0 5122 0 9178 CA 71 08 CAL 2 0038 0 7653 1 2049 CA HB2 HB3 OG 8 HB2 HALI 6328 1 7901 1 1839 CB OB 9 HB3 H_ALI 6328 0 3109 2 1235 CB OB 10 OB PSEUD 6328 1 0505 1 6537 11 OG O_HYD 3 4286 0 7774 1 2238 CB HG 12 HG H_OXY 3 1598 1 2840 2 0214 OG 13 6 C_BYL 9763 1 4377 0 0000 CA O N 140 O_BYL 1939 2 3868 0 0000 C 15 N_AMI 3 2963 1 5532 0 0000 C Ds File Formats Names of atoms located in the preceding or next residue are preceded by or respectively and positions that correspond to a 0 in the nor mal format are left blank The standard residue library also includes a statistical data base of chem ical shift values in proteins which was compiled by Daniel Braun on the basis of 26 proteins for which IH BC and PN assignments are avail able There are no random coil values in this table The first few lines of this data base are as follows CSTABLE 320 1 ALA N 144 123 04 3 67 133 90 130 40 117 60 113 70 3 ALA CA 184 52 59 2 19 57 30 55 60 48 96 47 15 4 ALA CB 182 18 78 1 96 24 20 22 70 15 70 14 50 5 ALA HN 159 8 16 0 76 10 14 9 29 6 73 6 19 6 ALA HA 169 4 32 0 53 6 16 5 24 3 54 2 94 7 ALA OB 167 1 36 0 24 erg oF 1 67 1 01 0 02 The number after the heading CSTABLE denotes the number of atoms for which chemical shift information is available For each such atom one line of data
67. 0 400 flatsteps n n 50 100 tf watsoncrick write aco Es Commands Performs a standard variable target function minimization G ntert et al 1991a starting at minimization level L and ending at minimization level Lp At each of the lower levels i e those below Ly N minimization steps are performed the minimization is stopped if n steps failed to decrease the target function by at least 1 and the steric repulsion is considered only for the heavy atoms The weight for steric lower limits is 0 2 At minimization level L three times N minimization steps are per formed the minimization is stopped if n steps failed to decrease the tar get function by at least 1 and the steric repulsion is considered for all atoms The weight for steric lower limits is 0 2 for the first N minimiza tion steps then it is increased to 0 6 for the following N gt minimization steps and to 2 0 for the final N minimization steps If the option tf and none of the other parameters is given the final tar get function value is calculated without performing any minimization strand1 residue range strand2 residue range planar Creates restraints to enforce standard Watson Crick type pase pairing between two antiparallel DNA or RNA strands The two strands must have the same length and continuous numbering Optionally additional planarity restraints can be added that restrain the interstrand C1 C1 d
68. 0 if the chemical shift is unknown Number of the diastereotopic partner of atom n If atom n has no ste reopartner the function returns 0 If the atom and its stereopartner are stereospecifically assigned see command atom stereo the function returns the number of the diastereotopic partner with a negative sign Target function value obtained from the most recent target function eval uation If the current structure was read from a file that contained the tar get function value in its header and if the target function was not re evaulated after reading the file then the value from the file header is re turned Target function value of structure n Length of the time step used in the last integration step of a previous md command Target function value of the current structure obtained by evaluation of the target function with the current constraints weights etc Minimal local target function value in the most recent grid search Maximal local target function value in the most recent grid search Local target function value of residue n in the current structure obtained by evaluation of the target function with the current constraints weights etc Tolerance for the coupling constant value of the n th coupling constant restraint Residue range Atom selection Selections The DYANA command groups atom angle peak distance and structure apply to sets of selected atoms angles peaks distance con straints and
69. 017 HB ILE WAS 5 deviations larger than tolerance All these peaks should be checked and modified before running NOAH If the assignment is correct the peak position should be shifted to the in tersection point of both proton shifts A typical NOAH script NOAH dya can be found in the noah exam ple directory of the DYANA distribution package dyanalib prot_nam er2 read seq prot_nam seq read aco prot_nam aco tolerance 0 01 0 01 0 3 tol_una 0 01 0 01 0 3 tol_transp 0 03 0 03 0 6 seed 3771 info normal nproc 6 rmsd_range 3 37 ssa atoms stereo list noah num 24 peak_nam er2_h2o_na er2_d2o_na rmsd rmsd_range protein prot_nam 31 Tutorial DA Analyzing NOAH output 32 proton_nam h20 d20 addupl ssbond addlol ssbond peak_ref er2_h2o er2_d2o calibrate First the protein sequence and the predefined angle constraint files are loaded Then the NOAH variables tolerance tol_una and tol_transp are set The variable tol_transp is only used for 3D spectra to check for transposed peaks If the script will run on a multi processor machine the line nproc 6 may be uncommented If some stereo specific assign ments are available they may be included into a macro called ssa dya and read before the noah macro is started The noah macro will perform 24 cycles num 24 and assign the two unassigned peak lists er2_h20_na peaks and er2_d20_na peaks T
70. 1 1355 9 HB3 HAL 1 6404 0 3509 2 1071 10 OB PSEUD 1 6404 1 0801 1 6213 11 SG S OXY 3 8460 0 8430 1 2654 12C CBYL 1 9334 1 4526 0 0000 130 BYL 1 1232 2 3781 0 0000 14N NM 3 2494 1 6072 0 0000 GLN RESIDUE GLN 6 23 8 22 OMEGA 2 10 0000 2 1 3 2 PHI 0 0 0 0000 1 3 5 3 CHI1 3 1 3500 3 5 7 4 CHI2 3 1 3500 5 Tai 5 CHI3 0 0 0 0000 7 11 15 6 PSI 0O 0 0 0000 3 5 21 C CBYL 0 6824 1 1357 0 0000 20 OBYL 0 1723 2 2550 0 0000 3 N_AM 0 0000 0 0000 0 0000 AHN HAMI 0 4226 0 9063 0 0000 5 CA CAL 1 4530 0 0000 0 0000 6 HA HAL 1 7427 0 5017 0 9233 7CB CAL 2 0013 0 7874 1 1917 8 HB2 HAL 1 6341 1 8130 1 1545 9 HB3 HAL 1 6341 0 3510 2 1206 142 NSNONNUWUWWHR HH 2000000o0o0ororoo ht ooooooooooo0ooo H H DONDONTDOCWMWDODHDOB Js DANGU GOOD ND sad AH RR H a oooooooorowoooo ht 0 30000 009000 0 0 000 H OORPONDOOODVDVONSOPOWONNOO jah DONO GO DG PRP oP WWA NON NOW Ww PRR PNARPNA m H H oooooooHhroNDoooo HH oOoO000000000000 H DI OOO ON OO O2 DDUSONSNUUWWH mr OOPFOOOOVONSOWMOO HH NNN IO II COWDWOHDOKBROWO oOVOoNOUOO R N oororoooo TOOTOO O COO BRR Ms File Formats 0 B PSEUD 1 6341 1 0820 1 6375 0 0 0 0 0 1 CG CAL 349913 0 7874 1 1917 7 12 13 15 0 2 HG2 HAL 3 8985 0 2382 1 2290 11 0 0 0 14 3 HG3 HAL 3 8985 1 2239 0 2629 11 0 0 0 14 4 06 PSEUD 3 8985 0 4928 0 7459 0 0 0 0 0 5 D C BYL 4 07
71. 1370 0000 0659 0000 1209 3784 1036 7356 7010 2498 9754 8690 1745 0716 2897 12357 6047 15778 o oO Wa gt 00 Ce 4817 7976 0167 5259 4216 4568 2157 8362 2779 5099 1963 1027 337 31 4019 7912 5965 0000 0659 0000 1209 2881 4402 3338 2250 2471 2464 0063 ooooooo000 00 00 00 00 00 00 00 00 00 oo wN o 00 00 00 00 Dj 4 4 3 ar 200 074 509 697 704 240 098 909 144 NONNNNWWWNHNNNN DNDN LY NN QO N NWUNNN 0215 5916 4863 8260 8907 3584 5959 1747 6228 4293 7403 6718 8894 9157 4100 7932 0000 9441 2311 0867 4736 4092 6979 07 384 Oo gt oo SIPUNDSONDOCOHKPOoonN OOOHr OOOHrOoOoOOOOHNHHHW o SEL Los 25 Di 2 0 0 H 11 13 19 22 30 37 sol T 7719 2842 3259 5214 1193 2135 2707 1041 6874 0000 8952 0000 5756 5780 6635 2499 9567 0000 0183 0000 8017 1364 0548 8814 4424 2228 7874 8681 OP 0730 00 wW RARA JOJJOUU N N 23 24 25 26 26 25 30 30 32 23 34 22 36 36 38 wW ww UD WWNNRFWE EE N oo N ORGE OOT U NO KD A ATO E Nad UU DN N N 23 23 25 25 27 27 N VOOO VvOoOoSsoPONDOOOD NO N Ww NNNN JOYO 39 ESC AOS WW WW WANN hop H Ww N N PO Rack N DOWDWDHDOBWODDTDCVODOHDOU
72. 197 Index BD write upl 119 write_all 119 198
73. 2 1065 7 0 0 0 10 0 QB PSEUD 1 6202 1 1030 1 6116 0 0 0 0 0 1 CG C VIN 3 5292 0 8520 1 2448 7 12 133 0 0 2 CD1 ARO 4 2978 1 4719 2 1507 11 16 17 0 0 3 D2 VIN 4 4485 0 2236 0 3267 11 14 15 0 0 4 CE3 CARO 4 2018 0 5582 0 8155 13 18 19 0 0 5 CE2 VIN 5 7342 0 5067 0 7404 13 16 20 0 0 6 NEL NAMI 5 6397 1 2900 1 8848 12 15 21 0 0 7 HD1 HARO 3 9116 2 0476 2 9918 12 0 0 0 0 8 HE3 HARO 3 1962 0 7957 ee dat 00 9 CZ3 CARO 5 3514 0094 4749 14 2 23 0 0 20 CZ2 CARO 6 8740 0 0475 0 0695 15 23 24 0 0 21 HE1 HAM 6 4722 1 6873 2 4654 16 0 0 0 0 22 HZ3 HARO 5 2167 6203 2 3675 19 0 0 0 23 CH2 CARO 6 6525 0 7332 0713 19 20 25 0 0 24 HZ2 HARO 7 8795 0 2851 0 4166 20 0 0 0 0 25 HH2 HARO 7 4966 1297 6404 23 0 0 0 0 26 C C_BYL 1 9258 4551 0 0000 5 27 23 0 0 270 OBYL 1 1108 2 3763 000 26 0 0 0 0 28 N LAM 3 2409 6166 0 0000 26 0 0 0 0 TYR RESIDUE TYR 6 28 3 27 OMEGA 2 10 0000 2 1 3 4 0 2 PHI O 0 0 0000 1 3 5 25 0 3 CHI1 Ca 5e A 4 CHI2 0 0 0 0000 5 7 14 15 25 5 CHIG 2 1 7500 17 19 24 25 25 6 PSI O 0 0 0000 3 5 26 28 0 C CBYL 0 6824 1 1357 0 0000 2 3 0 0 0 26 BYL 0 1723 2 2550 0 0000 1 0 0 0 0 3N NM 0 0000 0 0000 0 0000 4 5 0 0 4 HN HAMI 0 4226 0 9063 0 00 3 0 0 0 0 5 CA CAL 1 4530 0 0000 0 0000 3 6 7 26 0 6 HA HAL 1 7894 0 5104 0 90224 5 0 0 0 0 7 CB CAL 1 8747 0 6954 1 2959 5 8 9 14 0 8 HB2 HAL 1 4494 1 6989 1 3128 7 0 0 0 10 9 HB3 HAL 1 4494 0 1548 2 1414 7 0 0 0 10 10 OB PSEUD 1 4494 0 9269 1 727 o Gr
74. 2 3882 0 0000 19 0 0 0 21 N NMM 3 3043 5436 0 0000 19 0 0 0 0 GLU RESIDUE GLU 6 20 3 19 OMEGA 2 10 0000 2 1 3 4 0 2 PHI O 0 0000 1 3 5 148 0 3 CHIL 3 1 3500 3 5 wW TL 17 4 CHI2 3 1 3500 5 7 11 15 17 5 CHI3 0 0 0 0000 7 Jah 15 16 17 6 PSI 0 0 0 0000 3 5 18 20 0 C CBYL 0 6824 1 1357 0 0000 2 3 0 0 0 20 OBYL 0 1723 2 2550 0 0000 1 0 0 0 0 3 N LAM 0 0000 0 0000 0 0000 1 4 5 0 0 4 HN HAMI 0 4226 0 9063 0 0000 3 0 0 0 0 5 CA CAL 1 4530 0 0000 0 0000 3 6 7 18 0 6 HA HAL 1 7427 0 5017 0 9233 5 0 0 0 0 7CB CAL 2 0013 0 7874 1 1917 5 8 9 1 0 8 HB2 HAL 1 6341 1 8130 1 1545 7 0 0 0 10 9 HB3 HAL 1 6341 0 3510 2 1206 7 0 0 0 10 0 QB PSEUD 1 6341 1 0820 1 6375 0 0 0 0 0 1 CG C_AL 35313 0 7874 1 1917 7 12 13 15 0 2 HG2 HAL 3 8985 0 2382 1 2290 11 0 0 0 14 3 HG3 HAL 3 8985 1 2239 0 2629 11 0 0 0 14 4 06 PSEUD 3 8985 0 4928 0 7459 0 0 0 0 0 SD C BYL 4 0796 1 5749 2 3835 11 ie 17 0 0 6 OE1 OBYL 5 3227 1 6469 2 4925 15 0 0 0 0 7 OE2 O BYL 3 2432 2 0870 3 1585 15 0 0 0 0 8C CBYL 1 9838 4350 0 0000 5 19 20 0 0 90 BYL 1 2064 2 3882 0 0000 18 0 0 0 0 20 N NMM 3 3043 5436 0 0000 18 0 0 0 0 143 File Formats GLY HIS HIST HIST 144 SSS RESIDUE GLY 1 OMEGA 2 PHI 0 3 PSI 0 LG C_BYL 20 O_BYL 3N N_AM 4 HN H_AM 5 CA C_AL 6 HAl HAL 7 HA2 HAL 8 QA PSEUD cmo C_BYL 10 O O_BYL 11 N N_AM RESIDUE HIS OMEGA 2 PHI 0 3 CHIl 4 CHI2 0 5 PsI 0 E C_BYL
75. 20 O_BYL 3N N_AM 4 HN H AM 5 CA C_AL 6 HA H_AL 7 CB C_AL 8 HB2 HAL 9 HB3 HAL 0 OB PSEUD 1 CG C_VIN 2 NDI NAM 3 CD2 C_ARO 4 HD1 HAM 5 CET CARO 6 NE2 NAM 7 HD2 H_ARO 8 HE1 H_ARO IC C_BYL 20 O O_BYL 21 N N_AM RESIDUE HIST OMEGA 2 PHI 0 3 CHIL 1 4 CHI2 0 5 PSI 0 Cc C_BYL 20 O_BYL 3 N_AM 4 HN H AM 9 CA C_AL 6 HA H_AL 7 CB C_AL 8 HB2 HAL 9 HB3 HAL 0 OB PSEUD 1 CG C_VIN 2 NDI NAM 3 CD2 C_ARO 4 HD2 H_ARO 5 NE2 NAM 6 CE1 C_ARO 7 HE1 H_ARO 8 HE2 H_AM 9C C_BYL 20 O O_BYL 21 N_AM RESIDUE HIS 5 O Hs 01010 ds BW w WRPRRFRADUOA ds ds y 3 0000 0000 0000 1 2 0000 9063 0000 0 o 0 4284 3910 5208 0000 0000 3500 0000 0000 NO lt 0000 lt 0000 3500 lt 0000 lt 0000 oo 10 2 1357 2550 5343 5343 5343 900000060 000 0 Owe 0000000 J3J0UuU RP l fe O UE AA EN E ST OOONDOONDNONHHNHH oooooou_uwhr ee ae Kun oth Cho al oli O OOOONDNOOONHHNHH 0000 0000 0000 0000 0000 8762 8762 0000 0000 0000 0000 0000 0000 0000 0000 0000 9231 1699 0938 1019 5979 2388 2231 4368 9668 0126 9048 4399 6184 0000 0000 0000 14 19 0000 0000 0000 0000 0000 9231 1699 0938 1019 5979 2388 2231 4368 4399 9048 0126 6184 5438 0000 0000 0000 wwWU0NO 1M10WWNRRNA ds 19 11 12 HN WOONNWWNHNFPRPATODNANOOWWE 19 11 12
76. 21 HAM 3 4834 0 2913 0 5236 13 0 0 16 5 HD22 HAM 5 0670 0 0498 0 0896 13 0 0 0 16 6 QD2 PSEUD 4 2752 0 1208 0 2170 0 0 0 0 0 IC CBYL 1 9587 1 4440 0 0000 5 18 19 0 0 80 OBYL 1 1648 2 3835 0 0000 17 0 0 0 0 9 N LAM 3 2772 1 5756 0 0000 17 0 0 0 0 ASP RESIDUE ASP 6 17 3 16 OMEGA 2 10 0000 2 1 3 4 0 2 PHI O 0 0 0000 1 3 5 15 0 3 CHI1 3 1 3500 3 5 7 11 14 4 CHI2 0 0 0000 5 7 11 12 14 5 CHI32 2 200 7 11 13 14 14 6 PSI 0 0 0 0000 3 5 15 17 0 GC CBYL 0 6824 1 1357 0 0000 2 3 0 0 0 20 BYL 0 1723 2 2550 0 0000 1 0 0 0 3N NAM 0 0000 0 0000 0 0000 1 4 5 0 0 4 HN HAMI 0 4226 0 963 0 0000 3 0 0 0 0 5 CA CAL 1 4530 0 0000 0 0000 3 6 7 15 0 6 HA HAL 1 7394 0 5422 0 9011 5 0 0 0 0 7CB CAL 2 0038 0 6938 1 2474 5 8 9 1 0 8 HB2 HAL 1 6180 1 7132 409776 7 6 0 0 40 9 HB3 HAL 1 6190 0 1818 2 1294 7 0 0 0 10 10 0B PSEUD 1 6190 0 9475 1 7035 0 0 0 0 0 ince C BYL 3 5302 0 7444 1 3384 7 12 3 0 0 12 ODl O BYL 4 0951 1 2810 2 3031 11 0 0 0 0 13 002 O HYD 4 1537 0 1955 0 3514 11 14 0 0 14 HD2 H OXY 5 1416 0 2707 0 4866 13 0 0 0 0 15C CBYL 1 9587 4440 0 0000 5 16 17 0 0 160 OBYL 1 1648 2 3835 0 0000 15 0 0 0 0 17 N NM 3 2772 15756 0 0000 15 0 0 0 0 ASP RESIDUE ASP 5 16 3 15 1 OMEGA 2 10 0000 2 1 3 4 0 2 PHI 0 0 0 0000 1 3 5 14 0 3 CHI1 3 ss Aa S to ES 4 CHI2 0 0 0 0000 5 7 n R2 B 5 PSI 0 0 0 0000 3 5 14 16 0 10 GB 0 6824 1 1357 0 0000 2 3 0 0 0 20 BYL 0 1723 2 2550 0 000 1 0 0 0 0 141 File For
77. 23N NMM 3 3231 5208 0 0000 21 0 0 0 0 PHE RESIDUE PHE 5 271 3 26 OMEGA 2 10 0000 2 1 3 4 0 2 PHI O oO 0 0000 1 3 5 2355 0 3 CHI1 3 130 3 5 7 14 24 4 CHI2 0 0 0 0000 5 7 14 15 24 5 PSI 0 0 0 0000 3 5 25 27 0 C CBYL 0 6824 1 1357 0 0000 2 3 0 0 0 26 BYL 0 1723 2 2550 0 0000 1 0 0 0 0 3N NM 0 0000 0 0000 0 0000 1 4 5 0 0 4 HN HAMI 0 4226 0 9063 0 0000 3 0 0 0 0 5 CA CAL 1 4530 0 0000 0 0000 3 6 7 25 0 6 HA HAL 1 7765 0 5114 0 9066 5 0 0 0 0 7 CB CAL 1 9003 0 7053 1 2819 5 8 9 14 0 8 HB2 HAL 1 4836 1 7124 1 2948 7 0 0 0 10 9 HB3 HAL 1 4836 0 1770 2 1395 7 0 oO 0 10 10 B PSEUD 1 4836 0 9447 1 7171 0 0 0 0 0 11 D PSEUD 3 5529 0 8031 1 4598 0 0 0 0 13 147 File Formats PRO SER THR 148 Nu 12 QE PSEUD 13 OR PSEUD 14 CG C_VIN 15 CD1 C_ARO 16 HD1 H_ARO 17 CEL C_ARO 18 HE1 H_ARO 19 CZ C_ARO 20 HZ H_ARO 21 CE2 C_ARO 22 HE2 H_ARO 23 CD2 C_ARO 24 HD2 H_ARO 25 C C_BYL 26 0 O_BYL 27 N N_AM RESIDUE PRO 1 OMEGA T 2 PSI 0 LC C_BYL 20 O_BYL 3N N_AM 4 CD C_AL 5 CA C_AL 6 HA H_AL 7 CB AL 8 HB2 HAL 9 HB3 HAL 0 OB PSEUD 1 CG _ AL 2 HG2 HAL 3 HG3 HAL 4 QG PSEUD 5 HD2 HAL 6 HD3 HAL 7 QD PSEUD 8C C_BYL 90 O_BYL 20 N N_AM RESIDUE SER OMEGA 2 PHI 0 3 CHIT 4 CHI2 5 PSI 0 E C_BYL 20 O_BYL 3N N_AM 4 HN H_AM 5 CA C_AL 6 HA H_AL 7 CB C_AL 8 HB2 HAL 9 HB3 HAL 10 QB PSEUD 11 OG O_HYD 12 HG H_OXY 13 C C_BYL 140 O_BYL 15 N N_AM RESIDUE THR OMEGA
78. 25 HZ2 HAM 5 7617 3 4483 3 3787 23 0 0 0 27 26 HZ3 HAM BIER 241246 4 3351 237 0 O 0 27 27 02 PSEUD 6 2047 2 6920 3 7262 0 0 0 0 0 28 C C BYL 1 9587 1 4440 0 0000 5 29 30 0 0 290 OBYL 1 1648 2 3835 0 0000 28 0 0 0 0 30 N LAM 3 2772 1 5756 0 0000 28 0 0 0 0 MET RESIDUE MET 7 23 3 22 OMEGA 2 10 0000 2 1 3 4 0 2 PHI O eos 8 0000 T 3 Bx ar 0 3 CHIL ar 43600 5 5 m 11 20 4 CHI2 33 Led 05 AF CEE a 20 5 CHI3 3 1 0000 7 11 15 17 20 6 CHIA 3 200 11 15 17 18 20 7 PSI O 6 000000 3 57 21 28 0 G EB 0 6824 1 1357 0 0000 2 3 0 0 0 20 BYL 0 1723 2 2550 0 0000 1 0 0 0 0 3N LAM 0 0000 0 0000 0 0000 1 4 5 0 0 AHN HAMI 0 4226 0 9063 0 0000 3 0 0 0 0 5 CA CAL 1 4530 0 0000 0 0000 3 6 7 2 0 6 HA HAL 1 7457 0 5091 0 9188 5 0 0 0 0 7 CB CAL 159697 0 7431 lt 1 536 5e 8 CA 0 8 HB2 HAL 15896 1 1643 1231 E 70 30 9 HB3 HAL 1 5826 0202 2 187 7 010 0 QB PSEUD 1 5826 1 0134 1 68356 0 0 0 0 0 1CG CAL 3 4932 0 7637 120 7 12 13 15 0 2 HG2 HAL 3 8789 0 2557 1 2752 11 0 0 0 14 3 HG3 HAL 3 8789 1 2461 0 3722 11 0 0 0 14 4 0G PSEUD 3 8789 0 4952 0 8237 0 0 0 0 0 5 SD S RED 4 0596 1 6359 2 7211 11 17 0 0 0 6 QE PSEUD 6 1844 1 4722 2 4490 0 0 0 0 0 7 CE CAL 5 8250 1 4999 2 4950 15 18 19 20 0 8 HE HAL 6 3363 1 9959 3 3200 17 0 0 0 16 9 HE2 HAL 6 1084 0 4476 2 4720 17 0 0 0 16 20 HE3 HAL 6 1084 1 9731 1 5549 17 0 0 0 16 21 C CBYL 2 0013 4284 0 0000 5 22 23 0 0 220 BYL 1 9356 2 3910 0 0000 21 0 0 0
79. 50 0000 15 000 2 assign resid 4 and name N resid 4 and name CA resid 4 and name C resid 5 and name N 1 00000 60 0000 45 000 2 assign resid 4 and name N resid 4 and name CA resid 4 and name CB resid 4 and name CG 1 00000 60 0000 15 000 2 end set message on echo on end Each of the four atom selections for an angle constraint must match ex actly one atom In the above example the first assign statement con strains the p dihedral angle of residue 3 the second constrains y of residue 3 etc The allowed interval of a dihedral angle constraint is o Ad 6 Ad where and Ad are the second and third real number in the assign statement respectively Ds File Formats Scalar coupling constants Scalar coupling constant files specify values and optionally tolerance ranges and weighting factors for vicinal scalar coupling constants 1 ASP HA HB2 5 4 1 ASP HA HB3 4 1 2 GLU HA HB2 12 3 2 0 5 0 2 GLU HA HB3 4 1 2 0 5 0 2 GLU HN HA 5 1 0 3 CYSS HN HA 6 2 0 Each line specifies in this order the following data residue number res idue name first atom name second atom name value J of the coupling constant in Hertz tolerance AJ of the coupling constant default val ue 2 0 Hz and relative weight default value 1 0 The allowed interval of a coupling constant is J AS J AJ Orientation constraints Orientation constraint files specify values of
80. 541 CHI 154 468 CHI2 72 871 CHI3 174 205 CHI4 166 353 PSI 68 960 32 ARG PHI 39 181 CHI 153 419 CHI2 140 783 CHI3 56 936 CHI4 158 340 PSI 37 495 33 ILE PHI 64 011 CHI 81 197 CHI22 61 594 CHI21 135 938 CHI31 57 983 PSI 50 837 34 GLU PHI 54 036 CHI 98 207 CHI2 172 555 CHI3 5 197 PSI 41 856 35 ILE PHI 79 325 CHI 83 405 CHI22 171 980 CHI21 47 870 CHI31 145 152 PSI 16 608 36 ALA PHI 83 603 CHI 178 076 PSI 21 708 37 HIS PHI 113 998 CHI 110 027 CHI2 120 049 PSI 7 492 38 ALA PHI 114 971 CHI 166 685 PSI 12 535 39 LEU PHI 124 389 CHI 134 399 CHI2 39 785 CHI31 151 866 CHI32 61 206 PS 41 470 Output dihedral angle files from DYANA start with a comment line that indicates the final value of its target function Cartesian coordinates Cartesian coordinate files in DG format are used by DYANA for the input and output of conformations The format of the data lines is 6X A5 16 1X A5 3F11 4 corresponding to the atom name the residue number and name and the x y and z coordinates of the atom in For compatibility with other programs the first three lines are always com ment lines even if they do not start with further comment lines are not allowed Optionally the Cartesian atomic coordinates may be fol lowed by the covalent connectivities in this case the format is 6X A5 16 1X A5 3F11 4 416 On input Cartesian coordinates are only used to calculate all dihedral an gles the structure
81. 7480 8873 1513 9587 1648 2772 29 10 0000 3500 3500 3500 3500 9000 0000 oor ero 6824 12723 0000 4226 4530 7797 9888 6169 6169 6169 5188 8906 8906 8906 0546 6827 6827 6827 5845 9587 9587 9587 0907 1041 7789 4415 9587 1648 2772 30 10 0000 3500 3500 3500 3500 9000 0000 L 2 0000 9063 0000 oorerero 6824 1723 0000 4226 4530 0000 0000 005 0195 5960 4440 3835 5756 4805 8392 8507 430 1404 925 4120 9934 8015 7884 8326 7953 3524 4702 3800 R N HS0WwWHMN 0 15 3 2550 0000 9063 0000 4805 8392 8605 4400 1502 847 1742 2463 536 6863 7076 287 9974 694 6738 0943 384 5086 5293 4718 0006 4440 3835 5756 HH N 0arR J30WRANI 1357 2550 H nala EE EEE ana 15 ER DIA O r HPS0whr 15 19 OTTO 9222 1617 0656 0917 5787 2806 3849 7595 23553 4757 5369 3497 3920 3410 4253 2076 4331 0574 0000 0000 0000 LL 19 23 27 0000 0000 0000 0000 0000 9222 el 1 Ze ale i ils 0 0 D 2 3 2 23 2 4027 Lu B 3 gt lt 0000 lt 0000 lt 0000 1617 0798 1053 5926 1725 2551 2289 7420 3342 2516 2778 7647 3450 4289 9158 4723 5010 4151 4581 0000 0000 0000 0000 0000 Ss 415901000
82. 87 7 HOXI O 0 0 00 13 15 17 18 18 8 NUL 0 0 0 00 13 15 19 21 37 9 CHI 0 0 0 00 21 19 22 23 37 10 EPSI 0 0 0 00 11 13 38 39 0 103 CALI 0 9681 5 8551 2 5577 2 0 0 0 0 203 OEST 0 6348 4 7127 1 7719 1 3 0 0 0 3P PALI 0 4817 3 6875 2 2842 2 4 5 6 0 4 0P1 OBYL 1 7976 4 3635 2 3259 3 0 0 0 0 153 File Formats THY 154 ee 5 OP2 O BYL 6 05 OEST 7 C5 CAL 8 HS HAL 9 H5 HAL 0 05 PSEUD I gar E AL 2 H4 HAL 3 C3 CAL 4 H3 HAL 5 C2 CAL 6 H2 HAL 7 02 O HYD 8 HO2 H_OXY 9 Cl CAL 20 Hl HAL 21 04 O_EST 22 N9 NAM 23 C4 C_ARO 24 N3 N_AM 25 C2 C_ARO 26 N2 NAM 27 H21 H AM 28 H22 H_AM 29 02 PSEUD 30 N1 N_AM 31 Hl H_AM 32 C6 C_ARO 33 06 O_BYL 34 C5 CAR 35 N7 N_AM 36 C8 C_ARO 37 H8 H_ARO 38 03 O_EST 39 P P_AL RESIDUE THY 9 0 301014 MW NHO0Ot0o0 00H NOOO J004S UN NNNNNNNNNN ODANHDUPWNEO ZETA ALPHA BETA 0000000000 3 OP1 EE 3 3 WEA gt gt HWD DT aa In Ui 10 ol U un oO Q gt Q I D T T w x T Q w Q Pp PD gt gt UFFcCc A aa ak a ap EI na Eh aa na aga ia a AYE och NAN Jm ale Dp oO Q pa Q D tl Zz Zor w GRE AC Cee F fea ee ae HO Bee O ds 2 gt pe T w x E mcd WwW N Oooo OOo OOO OOo 2 oo N Wh 0167 5255 4216 4568 2197 8362 2779 5099 1963 1027 23131 4019 9936
83. 91 Index 192 delete 176 difima 176 directory 176 disconnect 176 distances 177 extract 177 help 177 insert 178 keep 178 link 178 list 178 pseudo 179 quit 179 read 179 remove 179 rename 179 retain 180 save 180 sort 180 type 180 writeaco 180 writeamber 181 writedco 181 writedg 181 writelongdco 181 writepdb 181 command 54 create 86 D dcostat 87 differences 87 dinucleotide 88 distance check 88 distance clear 89 distance comp 89 distance delete 89 distance keep 89 distance list 89 distance make 89 distance modify 90 distance scale 91 distance select 91 distance set 92 distance stat 92 distance unique 92 distance weight 92 DYANA functions acoviol 127 anam 127 Angle 127 angle 127 Atom 127 atom 127 calscale 127 cco 127 coord 127 derms 127 diastereotopic 127 dmax 127 dnam 128 drms 128 dval 128 ekin 128 ekmean 128 ekrms 128 element 128 emean 128 erms 128 heavyatom 128 iacod 128 iangle 128 iar 128 iatom 128 launit 128 ibond 129 iccoa 129 ida 129 idcoa 129 idord 129 idr 129 ifira 129 ifird 129 intervals 129 iprev 129 imum 129 istruct 129 Ida 129 libdir 129 maxang 129 maxcor 129 na 130 naco 130 nassign 130 nbond 130 neco 130 nconf 130 nd 130 ndcdis 130 ndco 130 ndcres 130 ndfree 130 nlevel 130 nlol 130 np_ass 130 np_corr 130 np_inc 130 np_new 130 np_out 131 np_wrg 131 npeaks 131 nplist 131 nr 131 nseldis 131 nstruct 131 numpro 131 nupl 131 pi 131 pseudoat
84. 96 1 5749 2 3835 11 16 17 0 0 6 OE1 BYL 5 2772 1 7032 2 5777 15 0 0 0 0 7 NE2 NAM 3 1391 2 0933 3 1681 15 18 19 0 0 8 HE21 HAM 2 1732 1 9506 2 9521 17 0 0 20 9 HE22 H_AM 3 3980 2 6258 3 9740 17 0 0 0 20 20 QE2 PSEUD 2 7856 2 2882 3 4631 0 0 0 0 0 21 CBYL 1 9838 14350 0 0000 5 22 23 0 0 220 OBYL 1 2064 2 3882 0 0000 21 0 0 0 23 N LAM 3 3043 5436 0 0000 21 0 0 0 0 GLU RESIDUE GLU 7 2 3 20 OMEGA 2 10 0000 2 1 3 4 0 2 PHI 0 0 0 0000 1 3 5 de 0 3 CHIL 3 S500 132 80 Fy JA 18 4 CHI2 3 3500 5 7 11 15 18 5 CHI3 O 0 0 0000 7 11 15 16 18 6 CHI42 2 4 0000 11 15 m 18 18 7 PSI 0 0 0 0000 3 5 19 21 0 C CB 0 6824 1 1357 0 0000 2 3 0 0 0 20 OBYL 0 1723 2 2550 0 0000 1 0 0 0 0 3 N LAM 0 0000 0 0000 0 0000 1 4 5 0 0 4 HN HAMI 0 4226 0 9063 0 0000 3 0 0 0 0 5 CA CAL 1 4530 0 0000 0 0000 3 6 7 19 0 6 HA HAL 1 7427 0 5017 0 9233 5 0 0 0 0 7CB CAL 2 0013 0 7874 1 1917 5 8 9 11 0 8 HB2 HAL 1 6341 1 8130 1 1545 7 0 0 0 10 9 HB3 HAL 1 6341 0 3510 2 1206 7 0 0 0 10 0 QB PSEUD 1 6341 1 0820 1 6375 0 0 0 0 0 1 CG C_AL 3 5313 0 7874 1 1917 7 12 13 15 0 2 HG2 HAL 3 8985 0 2382 1 2290 11 0 0 0 14 3 HG3 HAL 3 8985 1 2239 0 2629 11 0 0 0 14 4 06 PSEUD 3 8985 0 4928 0 7459 0 0 0 0 0 5CD C BYL 4 0796 1 5749 2 3835 11 16 17 0 0 6 Qui BYL 5 3043 1 6818 2 5453 15 0 0 0 7 OE2 O BYD 3 1835 2 0873 3 1571 15 18 0 o 0 8 HE2 HOXY 3 6219 2 5827 3 9070 17 0 0 0 0 9C CBYL 1 9838 4350 0 0000 5 20 21 0 0 200 OBYL 1 2064
85. A and C in amino acids will also be needed If the new residue type results from a slight modification of an existing residue type it is usually most Working with several molecules Ds Tutorial convenient to start from the coordinates of the existing residue type and to modify them Order the atoms such that their order is compatible with the tree structure of dihedral angles that will be defined i e such that the following two rules are fulfilled e A change of a dihedral angle must not affect the positions of the first second third or forth atom in any preceding dihedral angle defini tion e The set of atoms whose positions will be affected by a change of a dihedral angle consists of all atoms following the third atom in the dihedral angle definition up to the fifth last atom in the dihedral an gle definition or the end of the main chain for backbone dihedral an gles Convert the coordinates into the format of the library file for example with a text editor Add atom types connectivities and the information about diastereotopic partners Add the dihedral angle definitions to the new entry These two steps are best done using the library format in which connectivities and angle definitions are given by atom names rath er than by atom numbers see the option convert of the read lib com mand Make sure that the header line starting with RESIDUE is correct Add the new entry to a copy of the residue library file Test t
86. AL 5089 1 1747 0 8952 11 0 0 0 0 303 CAL 2 1963 0 6228 0 0000 11 15 14 38 0 4 43 HAL 3 1027 0 4293 0 5756 13 0 0 0 0 5 02 CAL 13731 F403 0 5780 19 16 17 13 0 6 H2 HAL 1 4019 6718 1 6635 IB 0 0 0 18 7 H2 HAL 7912 2 6897 0 2499 15 0 0 0 18 8 02 PSEUD 15965 2 1808 0 9567 0 0 0 0 0 9 C1 CAL 0 0000 4100 0 0000 21 20 22 15 0 20 HI HALI 0 0659 7932 21 0183 19 0 0 7 0 2104 OEST 0 0000 0 0000 0 0000 11 19 0 0 0 22 N9 NA 1 1209 9441 0 8017 19 23 36 0 0 23 C4 CARO 2 3784 2 2371 0 3436 22 24 34 0 0 24 N3 NA 2 8170 2 0867 0 9370 23 25 0 0 0 25 C2 CARO 4 1036 2 4736 1 0196 24 26 30 0 0 26 N2 NA 4 7356 2 4092 2 1892 25 27 28 0 0 27 H21 HA 5 7010 2 6979 2 2581 26 0 29 28 H22 HA 4 2498 2 0716 3 0078 26 0 0 0 29 29 02 PSEUD 4 9754 2 3848 2 6330 0 0 0 0 0 30 Nl NA 4 9152 2 9484 0 0482 25 31 32 0 0 31 Hl HA 5 8690 3 2001 0 2652 30 0 0 0 0 32 C6 CARO 4 4155 3 0749 1 2114 30 33 34 0 0 33 06 O BYL 5 1745 3 5092 2 0764 32 0 0 0 34 C5 CARO 3 0716 2 6978 1 4177 23 32 35 0 0 35 N7 NA 2 2897 2 7047 2 5414 34 36 0 0 0 36 08 CARO 1 1186 2 2402 2 1184 22 35 37 0 0 37 H8 HARO 0 2357 2 0985 2 7240 36 0 0 0 0 38 03 OEST 2 6047 0 9094 1 3466 13 39 0 0 0 39 P PAL 3 5778 2 1447 rear 38 0 0 RGUA RESIDUE RGUA 10 39 3 38 1 ZETA 0 0 0 00 L 2 2088 2 ALPHA 0 0 0 00 Pe By LI O 3 BETA 0 0 0 00 Ce eee ee 4 GAMMA 0 0 0 00 el ot Tal 432 F 5 DELTA 0 0 0 00 wie de 36 6 6 NU2 0 0 0 00 i AS 16 19
87. ARGE Same as large except that this setting can only be over ridden by another value written in capital letters FULL Same as full except that this setting can only be overrid den by another value written in capital letters This set ting is particularly useful for debugging macros in which the echo is suppressed Labels are not included in the echo but variable substitutions are State ments preceded by are only echoed if echo has the value full or FULL is a variable for error handling in macros If an error occurs within a mac ro the value of erract is executed as a command By default the exit command is executed i e the program returns to interactive input Errors that occur interactively are displayed and the program continues with the execution of the next statement set erract show quit In case of an error in a macro a listing of all glo bal variables is given and the program is stopped Such error handling can be useful if the program is used non interactively determines which messages are written to standard output and into the protocol file The possible settings are none No messages are written minimal A minimal set of messages is written in general a single line for each command that is executed normal The normal amount of messages is written full The full amount of messages is written debug The full amount of messages and additional undocu mented messages for debug pu
88. Calculating structures using torsion angle dynamics 14 Usage calculate file lt file gt struct lt n gt syntax file struct i 5 readdata file calc all struct write cor file all overview file Now the two parameters are available inside the macro under the names file and struct respectively The asterisk indicates that the value of the file parameter can be any character string whereas i restricts the struct parameter to have only integer values The struct parameter has a default value of 5 and there is no default value for the file parameter All the following calls of the calculate command are equivalent Calculate helix 5 Positional parameters calculate helix Default value for parameter struct calculate file helix struct 5 Named parameters calculate struct 5 file helix Any order of parameters calculate helix str 5 Abbreviated parameter name Lines at the beginning of the calculate macro that start with are comment lines used by the on line help system These are displayed when on line help is requested about the macro dyana gt help calculate calculate calculate a group of structures Usage calculate file lt file gt struct lt n gt The macro for the calculation of a structure by simulated annealing with molecular dynamics in torsion angle space torsion angle dynamics TAD is anneal The standard simulated annealing protocol that is used if the anneal macro is called witho
89. DA OS O S amp 00000000Uu N w ooooooooooooooOooONDOOOVOWOHOOOHOOON OO m ooooo0o oooo0o0o0 oooooo0000 BR Oooo coo coo coo cooc 5 90 0 0 5 0 5 OOo 2 0 0 O OGOOGO OTGA oo0oo0o0oo 155 File Formats LL LL2 LL5 LP LN PLM NLM LLM 156 RESIDUE LL 1 LB 0 1 91 PSEUD 2 02 PSEUD 3 03 PSEUD 4 Q1 PSEUD 502 PSEUD 6 03 PSEUD RESIDUE LL2 1 LB 0 1 Ql PSEUD 2 02 PSEUD 3 03 PSEUD 4 Q1 PSEUD 502 PSEUD 6 03 PSEUD RESIDUE LLS 1 LB 0 15 01 PSEUD 2 02 PSEUD 3 03 PSEUD 4 Q1 PSEUD 502 PSEUD 6 03 PSEUD RESIDUE LP 1 LB 0 1 Ql PSEUD 2 02 PSEUD 3 03 PSEUD ac PSEUD 5 0 PSEUD 6 N PSEUD RESIDUE LN 1 LB 0 1 Ql PSEUD 2 02 PSEUD 3 03 PSEUD 4 C3 PSEUD 5 03 PSEUD 6 P PSEUD RESIDUE PLM 1 LB 0 LE PSEUD 20 PSEUD 3N PSEUD 4 Q1 PSEUD 5 01 PSEUD 6 Q2 PSEUD 7 02 PSEUD 8 03 PSEUD RESIDUE NLM 1 LB 0 1 C3 PSEUD 2 03 PSEUD BP PSEUD 4 Q1 PSEUD 5 01 PSEUD 6 Q2 PSEUD 7 02 PSEUD 8 Q3 PSEUD RESIDUE LLM UI DIARIA AA EA OP BNHNNOOF Got OS Nhe eae GON Ne eee SE NPR Sas ooo Oreo oOo Om Pp 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 0000 10 0000 15 0000 0000 0000 0000 0000 0000 0000 0920 0000 0000 0000 0000 0000 3856 6824 1723 0000 9971 9971 0733 0733 0704 9681 6348 4817 3487 3487 0465 0465 9136 00 oorroo
90. DYANA Version 1 5 Users Manual Peter G ntert Christian Mumenthalerand Torsten Hermann Institut fur Molekula rbiologie und Biophysik Eidgen ssisc he Technische Hochsc hule C H 8093 Zurich Switzerland June 1998 1996 1998 by Institut fiir Molekularbiologie und Biophysik Eidgen ssische Technische Hochschule CH 8093 Ziirich Switzerland ontents DAVEE NON sea oo teed ar aed oat er Maran end PER a Rahs TOK AN Hisar a De O A ee LS A A eh Oe SN mee II Commands si A RR AEE Variables and Functions 2 121 Selections Aes be oe is ee eeu Da e 133 File PORDAS see ae es Hoe Le ed ee S7 COPIA ota dia nan an dial dat el E A LOO References amp Wake EG Ra ar 167 Introduction The main purpose of the program package DYANA Dynamics algo rithm for NMR applications is the calculation of three dimensional protein and nucleic acid structures on the basis of conformational re straints derived from the NMR experiments DYANA is the successor of DIANA G ntert et al 1991a Giintert amp W thrich 1991 It provides all functionality of DIANA and many extensions in particular a new effi cient structure calculation method torsion angle dynamics During the last years the steady increase in size of macromolecular structures that can now be solved by NMR has called for improved and more efficient structure calculation methods The program DIANA relied on conjugate gradient minimization of a variab
91. HB2 instead of HB2 HB3 as they are called in the standard residue library atom selection all atoms missing adapt d20 check One of the following actions is performed on the selected chemical shifts missing Lists all expected chemical shifts that are not present in the chemical shifts list If no atom selection is given only proton shifts are reported adapt Uses the positions of the assigned peaks to adapt the pro ton shifts The new shift is the average chemical shift po sition of all peaks assigned to the same proton d2o Deletes all NH shifts from the chemical shift list This command is useful for preparing an chemical shift list for the automatic NOESY spectrum assignment of a spec trum recorded in DO check Checks the current chemical shifts in two ways First the shifts are compared to the corresponding minimal and maximal values in the statistics of expected chemical shifts which is stored in the standard library file dya na lib Chemical shifts that lie outside of this range will be printed They are not necessarily wrong but should be checked with care In a second test this command uses the positions of the assigned peaks to check the chemical shifts for inconsistencies For every proton shift the me dian and the spread of the peak positions assigned to this proton are calculated and printed if the spread is larger than the corresponding tolerance value see system vari atom stereo
92. K Laskowski et al 1993 Optionally the background can be omitted nobackground and the residues with backbone angles that lie outside of the allowed regions are labeled n All available structure memories Creates n random structures in the structure memories i Actual seed number Creates a random structure in the structure memory 0 The target func tion value is automatically set to 0 A new seed number i for the random number generator may be specified file file o unknown erroriwarningiskip error append read ang read cco read cor Es Commands Reads an angle constraint file Constraints that involve unknown resi dues or angles can either cause an error a warning or can be skipped Optionally the angle constraints are appended to those already present file file T structure n 1 unknown erroriwarningiskip error Reads an angle file If there is a DYANA header in the file the target func tion value will be read from the header Optionally only the n th struc ture may be read from a multi conformer file Otherwise all m structures in a multi conformer file are read and stored as structures 1 m The first structure read will also be stored in the default structure memory 0 The presence of unknown residues or angles can either cause an error a warning or can be skipped file file o unknown erroriwarningiskip error append Reads a coupling
93. LAN Real function that returns the standard deviation of the n th fit parameter of the last linear least squares fit see plot subcommand fit Real function that returns the optimal value of the n th fit parameter of the last linear least squares fit see plot subcommand fit Real function that returns the probability that the x value of the last lin ear least squares fit would be exceeded by chance see plot subcommand fit Character function that returns the value of the environment variable with name s Integer function that returns the UNIX process identification number of the current process Character function that returns the value of the global variable with name s even if it is hidden by another variable with the same name or a blank string if no global variable with this name exists Character function that returns the value of the global variable with name s even if it is hidden by another variable with the same name or s if no global variable with the name s exists Integer function that returns the number of the character s Function that returns the argument x if n 0 or x otherwise The ar guments x and x can have any type Integer function that returns the starting position of the first occurence of the string s in the string s or zero if s does not occur as a substring in sy 1 Integer function that returns the starting position of the last occurence of the string s in the string s o
94. Lists all selected angles see chapter Selections The names of fixed an gles are enclosed in parentheses name angle selection on off clear Defines an external name for the selected angles Not more than one an gle may be selected per residue External angle names are used in place of the corresponding internal angle name from the residue library when reading input files and writing output files Initially or after the com mand angle rename clear external and internal angle names are iden angle set angstat clear angstat list angstat make anneal Es Commands tical With angle rename off renaming may be turned off temporarily until it is turned on again by angle rename on or by a new external angle name definition value r angle selection all angles All selected angles are set to the value r given in degrees Clears the angle statistics that is used to create redundant angle con straints in the REDAC strategy G ntert amp W thrich 1991 see angstat make Lists the current angle statistics see angstat make ang_cut 7 taken from variable ang_cut Adds the current structure to the angle statistics that is used to create re dundant angle constraints in the REDAC strategy Giintert amp Wiithrich 1991 see macro redac First the local target function value of every single residue is calculated by summing up all contributions to the target funct
95. NUONDVOODNIDVDOBRWOO N bo HH ooroooJounoooowvoo Ww OY 34 ja m Nu N N OWUWUWNOONDOOONSOPOHOOOVOOOOUOO Ww N N UI OWO H H w N N ooooooooooooooooooooNDooovoovooHhHoooHoo w ooooooooooWooowo JowoooHhHooonoo HH KIDANG AA IA ANGADANI OO OO OO OO CPD OO DO 000500000005 NN HH HH oVo0o60000000056500065656 0000000000000 URA PL NL 30 C7 C_ALI 31 H71 HALI 32 H72 HALI 33 H73 HALI 34 07 PSEUD 35 C6 C_ARO 36 H6 H_ARO 37 03 O_EST 38 P P_ALI RESIDUE URA 1 ZETA 0 2 ALPHA 0 3 BETA 0 4 GAMMA 0 5 DELTA 0 6 NU2 0 7 HOXI 0 8 NUl 0 SACHE 0 10 EPSI 0 1c3 C_AL 2 03 O_EST 32 PAL 4 OP1 O_BYL 5 OP2 O_BYL 6 05 O_EST TES AL 8 H5 HAL 9 H5 H_AL 0 05 PSEUD 1cC4 C_AL 2 H4 HAL 3 C3 C_AL 4 H3 HAL 5 C2 CAL 6 H2 HAL 7 02 O HYD 8 HO2 H_OXY 9 Cl C_AL 20 H1 HAL 21 04 EST 22 N1 AM 23 C2 C_ARO 24 02 O_BYL 25 N3 AM 26 H3 H_AM 27 C4 C_ARO 28 04 O_BYL 29 65 C_AR 30 H5 C_AL 31 C6 C_ARO 32 H6 H_ARO 33 03 O_EST 34 P P_AL RESIDUE PL LB 0 C PSEUD 20 PSEUD 3N PSEUD 4 Q1 PSEUD 5 Q2 PSEUD 6 03 PSEUD RESIDUE NL 1 LB 0 1 C3 PSEUD 2 03 PSEUD 3 P PSEUD 4 Q1 PSEUD 502 PSEUD 6 03 PSEUD jah oooooooo0o000000 WNE 8531 8409 5641 0420 8157 0061 0464 6047 5778 Oyo Wo A 00 or 4817 7976 0167 3255 4216 4568 2151 8362 2779 5099 963 1027 3731
96. OO SOOOWOOOVOVOSOUOO ooovooo r roooooo000000 pa U N N Ms File Formats 20 HE HA 6 0160 1 0017 1 6515 19 0 0 0 0 21 CZ C VIN 6 3367 2 0321 3 3506 19 22 26 0 0 22 NH1 NAMO 7 6712 1 9691 3 2468 21 23 24 0 0 23 HH11 HA 8 2477 2 3929 3 9454 22 0 0 0 25 24 HH12 HA 8 0907 1 4983 2 4706 22 0 0 0 25 25 QH1 PSEUD 8 1692 1 9456 3 2080 0 0 0 0 0 26 NH2 N AMO 5 7747 2 6630 4 3906 21 27 28 0 0 27 HH21 HA 6 3512 3 0868 5 0893 26 0 0 0 29 28 HH22 HA 4 7788 2 7101 4 4681 26 0 0 0 29 29 QH2 PSEUD 5 5650 2 8985 4 7787 0 0 0 0 0 30C CBYL 1 9838 1 4350 0 0000 5 31 32 0 0 310 OBYL 1 2064 2 3881 0 0000 30 0 0 0 0 32 N LA 3 3043 1 5436 0 0000 30 0 0 0 0 ASN RESIDUE ASN 5 19 3 18 OMEGA 2 10 0000 2 1 3 4 0 2 PHI O Os 0 0000 AT 38 ar ag 3 CHI1 3 1 3500 3 6 7 dp ie 4 CHI2 O 0 0000 5 7 11 I2 16 5 PSI O 0 0 0000 3 5 17 19 0 C CBYL 0 6824 1 1357 0 0000 2 3 0 0 0 20 BYL 0 1723 2 2550 0 000 1 0 0 0 0 3N LAM 0 0000 0 0000 0 0000 1 4 5 0 0 4 HN H AMI 0 4226 0 9063 0 0000 3 0 0 0 0 5 CA CAL 1 4530 0 0000 0 0000 3 6 7 17 0 6 HA HAL 1 7394 0 5422 0 9011 5 0 0 0 0 7CB CAL 2 0038 0 6938 1 2474 5 8 9 1 0 8 HB2 HAL 1 6375 1 7196 1 2891 7 0 0 0 10 9 HB3 HAL 1 6375 0 1881 2 1409 7 0 0 10 0 QB PSEUD 1 6375 0 9538 1 7150 0 0 0 0 0 1 CG C BYL 3 5338 0 6925 1 2451 7 12 133 0 0 2 61 BYL 4 1821 1 1949 2 1483 11 0 0 0 0 3 ND2 NGAM 4 0726 0 1015 0 1825 11 14 15 0 0 a HD
97. RESIDUE CYT 9 36 3 35 1 ZETA 0 0 0 00 i a 3 eS a 2 ALPHA 0 0 0 00 ee ee o 3 BETA 0 0 0 0 3o oeh A 4 GAMMA 0 0 0 00 6 7 i 13 0 5 DELTA 0 0 0 0000 7 11 13 35 0 6 NU2 O 0 0 0000 11 13 15 19 34 7 NUL 0 0 0 0000 13 15 19 21 34 8 CHI O 0 0 0000 21 19 22 23 34 151 File Formats RCYT 152 N 000 J0UiH4swNROwOoO SINN BPBWMNHVO NNN wNH WWWWNHNNNN DY WNROw 00101 ww Oe 36 EPSI G3 03 OP1 OP2 05 Es H5 Qs T gt DH P RESIDUE 90 30014 UYNROwOoO 004 WN RO 0 J004awNnNRA NMNNNNNNN ND AUDUBWNHERO ZETA ALPHA BETA GAMMA DELTA NU2 HOXI NUl CHI EPSI 38 03 P OP1 OP2 05 Es H5 H5 Qs ca HA c3 H3 Cet H2 O2 HO2 Cl Hl 04 N1 C2 02 N3 C4 N4 0 EE AOS pa Ara GO m Aa oe PP PPP PS a oO Y ma al D Q D m ES KE ETIC EDAS EIEE UF EH TTE U CF dy dede E Ela BeBe Pie t BAN HHO RCYT 0000000000 3 ele eink D DP HWP YP Gree MK E QE 2 55 oO Aa R Omprrrrrhp SK PPE KO Q Q O Q ka b sl O Noe 3 CLE vs HE A Q O z WN ja 00000200000 Ooo a S S N W N 0 0000 9681 6348 4817 7976 0167 5255 4216 4568 2157 8362 2779 5099 1963 1027 373 4019 7912 5965 0000 0659 0000 1209 2 2 3 gt 4 D 4 4 I 0 0
98. RP 000d0Ro0o0rJOJ3 J00 NN 27 AS 19 23 24 29 NN ANI NG DING ANGINA E OA a rO aT A Ww NN NN m m N N oOoWOOOOOOOONSOoOOONDOOO WO DDNDNMND OoVVNNNNOO H N N N E oOoOVvOoOoOooOPOOOOOOONSOOONDOOOVD ON O PO Ww m m N N ooosooooHhrooo Nooouooowo ja N N N gt oOoOVOOOWOOOHOOONOOOWOOODSONOUMO CO NSIOoUuoo oooooooNDoooovooovoooho N m m o00o0o0o000o0o0ow4ooovooodnunoooreJoooso ooooo NN OONDNDOOOVDVOVOOPPROOOOOOOOO0OO0O NN DADYA ooooo Ms File Formats 6 HA HAL 1 7797 0 4805 0 9222 5 0 0 0 0 7CB CAL 1 9888 0 8392 1 1617 5 8 9 1 0 8 HB2 HAL 1 6169 1 8605 1 0798 7 0 0 0 10 9 HB3 HAL 1 6169 0 4400 2 1053 7 0 0 0 10 0 QB PSEUD 1 6169 1 1502 1 5926 0 0 0 0 0 1 CG CAL 3 5188 0 8471 1 1725 7 12 13 15 0 2 HG2 HAL 3 8906 0 1742 1 2551 11 0 0 0 14 3 HG3 HAL 3 8906 1 2463 0 2289 11 0 0 0 14 4 06 PSEUD 3 8906 0 5361 0 7420 0 0 0 0 0 5 CD CAL 4 0546 1 6863 2 3342 11 16 17 19 0 6 HD2 HAL 306827 2 7076 2 2516 15 0 0 0 18 7 HD3 HAL 3 6827 1 2871 3 2778 15 0 0 0 28 8 QD PSEUD 3 6827 1 9974 2 7647 0 0 0 0 0 ICE CAL 5 5845 1 6941 2 3450 15 20 21 23 20 HE2 HAL 5 9587 0 6738 2 4289 19 0 0 0 22 21 HE3 HAL 5 9587 2 0943 1 4027 19 0 0 0 22 22 QE PSEUD 5 9587 1 3841 1 9158 0 0 0 0 0 23 NZ NAMO 6 0907 2 5086 3 4723 19 24 25 26 0 24 HZ1 HAM 7 0907 2 5032 3 4648 23 0 0 0 27
99. Tali GE Tiba LL ul LL Ely EL TE EE Eb EN linkers CYT 101 THY CYT THY ADE ADE THY GUA GUA CYT THY THY THY CYT 2nd DNA strand NL 150 LL2 LL2 LL2 LL2 LL2 LL2 LL2 E2 GE EZ TE ERZ TEL TP linkers ALA 200 LEU protein NOAH Mumenthaler amp Braun 1995 Mumenthaler et al 1997 is an al gorithm for the automatic assignment of 2D and 3D NOESY spactra In iterative cycles new possible assignments are identified and tested through a structure calculation Alternative assignment possibilities for individual peaks are included simultaneously in these calculations and peaks are unambiguously assigned after the structure calculation only if the distance constraint from one of the assignment possibilities was clearly less violated in the structures NOAH was implemented in DYANA at two different levels First new commands were implemented for NOAH specific tasks assign create filter keep reliability write ass and new variables were introduced for NOAH specific parameters tolerance tol_una tol_transp etc Checking input for NOAH Ds Tutorial Second a noah macro was written which contains the NOAH schedule Two additional macros noahmin and noahanneal are called by the noah macro itself and should not be modified by the user The following sections give some practical advice on the use of NOAH DYANA The mandatory input is a peak list containing peak positions and vol umes in XEASY format and a list of chemi
100. _cut was posi tive in the previous cycle structures will be calculated using N and N 111 Commands reliability rmsd 112 minimization steps at intermediate levels and at the final level respec tively redac er2 schedule 1 0 1 0 0 4 0 0 0 0 50 In the first cycle 50 structures are calculated with the original constraints and angle con straints are generated with ang_cut 1 0 After this new structures are calculated three times using the previously generated constraints No new angle constraints are generated the third time Finally the structures are minimized on the last level In this cycle too no angle con straints are generated dist 1 0 Calculates the reliability distance RD Mumenthaler amp Braun 1995 of all unambiguously assigned peaks using the given tolerance range and the structures in the structure memory If dist is specified the number of assigned peaks with a RD above dist will be written After this com mand all peaks that cannot be explained with the current structures and the given tolerance range see System Variables are selected reliability 1 0 Calculate the reliability distance of all peaks write peaks incomp peaks selected Write all selected peaks into the peak file in comp peaks range residue range all residues segment n 3 Calculates pair wise root mean square deviation RMSD between all pairs of selected structures McLachlan 1979 for
101. a structure or a structure cluster joins another cluster This RMSD is the minimal RMSD that any of the structures in the first cluster have to any of the structures in the second cluster Cur rently up to 20 structures can be analyzed list string 1 2 3 w1 w 5 0 w2 w 10 0 Creates upper limit distance constraints of obsdis dpseud A dpseud S the pseudo atom correction if appropriate on the basis of the three dif ferent assignment lists of NOAH see command filter for information on the NOAH peak lists The parameter list is a string containing the num bers of the lists that should be considered list 1 Unambiguous assign ment list UAL list 2 Ambiguous assignment list AAL list 3 Test assignment list TAL Constraints in the UAL are weighted with the factor w by default or with the factor w if they are unambiguous based on the chemical shift alone see command assign Constraints from the AAL are weighted with 1 0 and constraints from the TAL with 1 0 Nass where N g is the number of possible assignments a peak has in the assign command The AAL only exists after the filter command and the TAL only exists after the assign command dcostat differences Es Commands file name dcostat ps Produces a graphics output file with the given name a GRAF file if the extension is grf a MIF file if the extension is mif or a Postscript file otherwise containing two p
102. additional parameters that do not match with one of the formats cakk Has the same meaning as except that additional pa rameters must not contain an sign Formats must not contain blanks A type can be one of the following Oi Or Any character string Integer expression Real expression A INCLAN lt i lt u lt r lt u Integer or real expression in the given range ii Integer range i e one of the following m a single integer expression MN two integer expressions Ma using the default value for n N using the default value for m name name List of mutually exclusive literals f extension Filename that will be extended with the given exten sion if necessary extension can also be name to denote the value of a preceding parameter command read_file syntax format asc bin file f format weight r 1 0 The command read _file has three parameters The first parameter format is required and can either be asc or bin the second parameter file is also required and is a filename that will be given the extension asc or bin depending on the chosen format and the third parameter weight is an optional real number with default value 1 0 end read_file asc test Positional parameters and default value for weight Equivalent to setting format asc file test asc and weight 1 0 read_file file test format asc weight 2 0 Named parameters in any order s
103. addlol file exit cycle num_cyc entry cycle 1 calibrate Automatically assigns the peak lists given in the array peak_nam with corresponding proton lists in the array proton_nam inn NOAH cycles If the peak lists do not contain a line with DYANAFORMAT the for mat of every peak list must be given in the string plformat The name given by protein will be used for output files namely the final overview and coordinate files Optionally the minimizer macro may be changed and reference peak lists peak_ref containing the correct assignment may be given Agreement between NOAH assignment and reference as noahanneal noahmin overview A Commands signment will be indicated in the final noah grf file The string given in options is used as parameters for the command assign e g op tions transposed 0 should be used if the transposed peaks should be checked in 3D peak lists Upper and lower limits distance constraint files may be added to each NOAH structure calculation e g constraints for known disulfide bonds with the parameters addupl and addlol The parameters entry and exit indicate at which cycle of the n cycles NOAH actually starts and ends which allows to split a given NOAH cal culation into several smaller jobs The parameter entry can only be used if all files necessary for the given cycle are present in the current direc tory i e structures assignment files etc The peaks are
104. ain the ways by which commands can write output to the standard output device in the following simply called screen and or to disk files by using the protocol mechanism or output redirection The concepts of this section do not apply to output that is written to ex plicitly named disk files by specific output commands All output has an importance level and only output that is important enough is actually written The definition of what is important enough is given by the special variable info that can in its simple form take one of five information level values none no output at all except for error messages minimal minimal output in general a one line confirmation normal the normal amount of output full detailed output debug additional undocumented debugging output The output can be duplicated into a protocol file In fact different info values might be used for output to the screen and to the protocol file In this case the info value consists of two simple info values separated by a comma A protocol file is written if the protocol variable is defined and has a non blank value that is the name of the protocol file If the file does not exist when the protocol variable is set to the corresponding name it is created otherwise the output is appended to an existing pro Output redirection alias A INCLAN tocol file protocol logfile Open protocol file logfile info minimal full Minimal scr
105. alue 102 Maximal proton acceptor distance for a hydrogen bond Initial value 2 4 A Maximal angle between the donor proton bond and the line connecting acceptor and donor for a hydrogen bond Initial value 35 Minimization level L G ntert et al 1991 Only distance constraints between atoms not more than L residues apart are considered in the tar get function L 0 only intraresidual L 1 intraresidual and sequential constraints etc Initial value number of residues 1 e use all distance constraints maxamb nstep obsdis ori_axial ori_rhombic seed soft_aco soft_cco soft_lol Ms Variables and Functions NOAH variable Maximum number of possible assignments a peak can have to be taken into the test assignment list Initial value 2 Number of steps n per dihedral angle in grid searches The grid search es will run over the n angle values 0 A 2A n 1 A where A 27 n Initial value 36 i e A 10 Maximal distance for which an NOE can be observed Used by automat ic calibration function calscale and for automatic assignment Initial value 5 0 A Axial component D of the tensor that relates residual dipolar couplings to the orientation of the corresponding chemical bond 8 8 6 D 3cos 0 1 D sin cos26 7 6 6 6 is the residual dipolar coupling as a function of the polar angles O and 6 of the chemical bond with respect to the principa
106. amino acid residue and LN to link a generic linker to a following nucleotide residue There is an additional set of linker residues PLM NLM LLM LLM2 LLM5 LPM and LNM with three instead of one rotatable angle that should only be used for torsion angle dynamics calculations with inertia tensors derived di rectly from atomic masses and positions LGLY is a linker residue with the geometry of GLY but only containing pseudo atoms In addition to the standard residue library dyana lib which is based on the ECEPP 2 force field Momany et al 1975 N methy et al 1983 a residue library amber lib that employs the standard geometry of the AMBER force field Cornell et al 1995 is also provided The names of atoms and dihedral angles are the same in both libraries The program DYANA supports an alternative format for residue library entries that uses atom names instead of numbers to define dihedral an gles covalent connectivities and pseudo atoms This format is particu larly useful in the process of creating manually a new or modified library entry As an example an alternative entry for SER is given which is equivalent to the one in the standard library RESIDUE SER 5 15 3 14 1 OMEGA 2 10 0000 0O ae N HN 2 PHI 0 0 0 0000 C N BAG 3 CHIl 1 3 1 3500 N CA CB OG HG 4 CHI2 1 3 0 3000 CA CB OG HG HG 3 es 0 0 0 0000 N CA N 176 C_BYL 0 6824 1 19353 0 0000 0 N 20 O_BYL 0 1723 2 2550 0 0000 C 3 N_AM 0 0000
107. an have the following values normal Coordinates are specified explicitly on the command line rotate textsize weight x0 yO x1 y1 XO YO X1 Y1 A INCLAN list Coordinates are taken from vector expressions and the corresponding command is applied to all points in the list The input mode is automatically set to list when a graphics file with list data is read Initial value normal defines the direction in which text is written and can have the following values off Text is written horizontally from left to right on Text is written vertically from bottom to top Initial value off defines the font size in points 1 pt 0 353 mm Initial value 12 defines a font property with the possible values regular Regular not bold bold Bold The Symbol font is only available as regular Initial value regular define the positions of the two reference points xo yg and x y in the standard coordinate system The standard coordinate system has its origin in the center of an A4 sheet and uses points 1 pt 0 353 mm to measure distances in both dimensions The x axis points to the right and the y axis points up Initial values x 250 y 375 x 250 y 375 define the positions of the two reference points Ag Y and X Y in the user coordinate system All positions and distances are measured in the user coordinate system except for linewidth text size mark size and das
108. angle specifications on the command line The asterisk represents all atoms or angles which are not explicitly given macro List the contents of the macro file s that match the given macro specifi cation A macro specification is a macro file name possibly containing wildcard characters but excluding the extension cfm file Write an angle constraint output file in the format used by the program DYANA see above This command can only be used with angle con straint files writeamber writedco writedg writelongdco writepdb macro Istring A COFIMA file Write a Cartesian coordinate output file in AMBER format the format used by the molecular dynamics program AMBER This command can only be used with coordinate files file Write a distance constraint output file in the format used by the program DYANA The residue name and number of the first atom of the constraints are not repeated if they are the same as for the previous constraint This command can only be used with distance constraint files file Write a Cartesian coordinate output file in DG format the format used for example by the program DYANA This command can only be used with coordinate files file Write a distance constraint output file in the format used by the program DYANA The residue name and number of the first atom of all constraints are
109. angle with corners x4 Y1 X1 Y2 X Y1 and x3 yo This command can also be used in list mode where xj y1 x2 and yo are four vector expressions In list mode the command can also be used without parameters In this case a rectangle with corners x _ 72 0 x x 1 2 0 x _ x 2 y and x x 1 2 ie a histogram bar is drawn for each point x yi in the list columns for the first and last point x _ and x are replaced by the minimal and maximal x values Kg and A respec tively axis fi exact list mode performs scaling of the given axis X or y on the basis of the vector ex pressions f Scaling sets the coordinates of the reference points in the user coordinate system X and X for the x axis and Y and Y for the y axis such that they include all values of the vector expressions f If the option exact is present then the new coordinates of the reference points will correspond exactly to the minimum and maximum of the vec tor expressions f otherwise a small margin will be added in order to avoid that points lie exactly on the boundary set shape shape spline spline square A INCLAN parameterzvalue sets one or several plot parameters to the given values The keyword set 1s optional X1 Y1 X2 Y2 X3 Y3 KAYA X5 Y5 X6Y6 normal mode draws a shape enclosed by a closed B zier spline curve that is defined by
110. ar groups of ASP and GLU by 180 such that there is a best fit between all selected structures This command does not affect the three dimen sional structure The change is limited to the nomenclature which results in a lower heavy atom RMSD 93 Commands forall graf grid aco 94 commands gt save parallel This macro performs a loop over all selected structures copies them into the structure memory 0 executes the user commands and copies the structure back to the structure memory Optionally the structures are saved as angle files with name dnnnmmm ang nnn denotes the current process number and mmm the structure number before they are copied back The calculation may be executed in parallel if the INCLAN vari able nproc has a value larger than 1 forall parallel vtfmin steps 100 800 angstat make Minimize all selected structures using the mac ro vtfmin with the given number of steps and in clude the resulting structures into the angle statistics file file replace Convert a graphics file into a postscript or MIF plot file If file has the extension grf nothing happens If file has the extension mif a MIF file with this name is produced from the corresponding graphics file with extension grf In all other cases a Postscript file is produced from the corresponding graphics file with extension grf If the option replace is set then the graphi
111. automatic assignment of NOESY spectra on the basis of known sequential resonance assignments peak positions and peak in tensities Mumenthaler er al 1997 Initial NOESY cross peaks assign ments derived from matching chemical shifts are subsequently refined in several cycles consisting of structure calculations using an error tolerant target function followed by an assessment of the possible peak assign ments in the light of the preliminary three dimensional structures ob tained This method has the potential to largely replace the manual method for NOESY assignment Given that currently the NOESY as signment and the collection of conformational constraints for a protein of 150 amino acid residues may require several months of manual work the far reaching consequences of the availability of a reliable au tomatic NOESY assignment method become evident Since the calculation of macromolecular three dimensional structures is a computationally intensive procedure it is important to make best pos sible use of the available computing resources With the advent of a va riety of parallel computers and distributed computing networks the optimization of our structure calculation software which was up to now geared to the requirements of vector supercomputers has turned towards optimal parallelization A structure calculation that consists of the inde pendent generation of many conformers has a high degree of inherent Ms Introduction para
112. automatically recognizes many UNIX computer systems The op tional parameter install directory denotes the directory where libraries macros and on line help files will be installed The default install direc tory is HOME lib where HOME is the value of the corresponding UNIX environment variable The install directory should be different from the directory in which the tar file was unpacked Options include d Use double precision 64 bit for real numbers default f Use the Fortran 90 compiler instead of Fortran 77 g Prepare executables for debugging h Print a summary of these options q Quick Compile without optimization S Use single precision 32 bit for real numbers t type Configure for a given computer type Possible types are listed in the file scripts identify which is used to deter mine automatically the type of the current computer sys tem The script configure assumes that the name of the directory where the program resides is of the form dyana version All parameters set by the configuration script are listed and stored in the file make config Exe cution of the configure script has no other effect than creating the file make config The program package is then built by the UNIx command make and installed in the directory install directory dyana 1 5 by make install The directory in which the tar file was unpacked can be removed after this step Executable shell scripts
113. cal shifts in XEASY format proton list NOAH can only yield good results if the resonance assign ment is complete i e if all or nearly all chemical shifts are assigned and if the chemical shifts given in the proton list agree with corresponding peak positions within a small tolerance range A in general 0 01 ppm in 2D and 0 02 ppm in 3D This is best achieved by manually assigning at least one NOESY peak for every proton shift e g by taking over the TOCSY peak list from the sequential assignment and overlaying it to the NOESY spectrum All assigned peaks in the input peak list will be re garded as safe NOAH will include them in every structure calculation and never delete or change them even if they give rise to large constraint violations Additional input will help NOAH to converge Dihedral angle constraints may be generated using coupling constants with the grid commands and known disulfide bridges should be included as upper and lower limit dis tance constraints use the DYANA commands ssbond and write upl to generate and save these distance constraints Before runing NOAH the input data should be checked for obvious in consistencies using some of the DYANA commands First no such warn ing messages should appear when loading a proton list dyana gt read prot kt WARNING Inconsistency for LYS 21 QB 1 617 HB2 1 618 HB3 1 580 WARNING Inconsistency for LYS 21 QG 1 774 HG2 1 424 HG3 1 741 C
114. calibrate A2 d 6 The logical xor operator is used in the second peaks select command to select all HN HN peaks except those that were already selected GRAF a part of INCLAN is a versatile tool to produce graphics both in Postscript and FrameMaker MIF format DYANA provides several commands to create standard plots For exam ple the following commands create a Ramachandran plot for the group of structures calculated in section Running DYANA at the beginning of this manual readdata helix read_all helix cor ramachandran rama ps The result is a Postscript file with name rama ps An additional com mand creates an equivalent output file rama mif in FrameMaker MIF format ramachandran rama mif 21 Tutorial DI 00 o ex o N Number of constraints gt o o 0 2 4 6 8 10 Range li jl S o Ww o 10 Number of constraints e 30 32 34 36 38 Sequence 38 RRRRIEIAHAL yy i TM donti i 1 dentivi 1 Mi dyn i i 2 AA donti i 2 aaa a _ Wit m degli i 3 don i 4 RMSD cluster analysis Residue range 29 39 N Ma A Q 0 01 02 03 04 05 RMSD 22 Two plots of the distribution of distance constraints as a function of their range i e the residue number difference and their residue numbers re spectively are created by the commands readdata helix dcostat dco that create a Postscript file
115. cards to match any number of characters and to match exactly one character If an angle declaration should apply only to part of the sequence the declaration and the corre sponding part of the sequence are enclosed in curly braces More than one angle declaration may follow the left brace and parts of the se quence enclosed in braces may be nested In the following example se quence all angles will be fixed at 180 by default all y angles except the third one will be fixed at 47 and all other dihedral angles are rotat able by default PSI 47 ARG 29 ARG PSI fr ARG ARG ILE GLU ILE ALA HIS ALA LEU 161 File Formats I Chemical shift list Chemical shift lists traditionally called proton lists follow the format used by the program XEASY Bartels ef al 1995 For each chemical shift the list contains a line with the following data in free format the atom number the chemical shift the error of the chemical shift current ly not used by DYANA the atom name and the residue number The atom number is referenced by peak assignments in peak lists see next section and can be different from the atom number in coordinate files Chemical shifts are measured in ppm and entries with a magnitude larger than 900 ppm are skipped An example containing proton ISN and BC chemical shifts follows 622 119 770 0 000 N 3 21 8 635 0 000 HN 3 22 52 530
116. ces and constraints that cannot be violat ed retains maximally one distance limit for each atom pair and introduces corrections for constraints with diastereotopic substituents for which stereospecific assignments are not available The result of this step are the modified upper distance limits in the file er2 upl the dihedral angle constraints in the file er2 aco and ste reospecific assignments The actual structure calculation is performed with the macro ANNEAL using torsion angle dynamics read seq name seq read sequence and initialize read upl name upl read upper distance limits read aco name aco read angle constraints ssbond 5 20 12 37 17 28 generate constraints for S S bonds seed 35621 random number generator seed nproc 4 number of processors calc_all 30 simulated annealing Building an INCLAN macro Ds Tutorial overview name structures 20 ang cor produce overview After reading the input data file constraints for the three disulfide bonds 5 20 12 37 and 17 28 are generated with the macro ssbond The ran dom number generator seed and if applicable the number of processors that can be used in parallel is set and 30 conformers are calculated with simulated annealing in torsion angle space using the standard annealing protocol i e the macro anneal For larger proteins it could be necessary to increase the number of time steps and or the number of conformers Finally the 20 best conformers are analyzed
117. covalent connectivities and the stan dard geometry The standard library dyana lib uses the standard ge ometry of the ECEPP 2 force field Momany et al 1975 N methy et al 1983 for all amino acid residue types The covalent geometry of the nu cleotides is based on the AMBER force field Cornell et al 1995 For reasons of compatibility with other programs the residue library used for DYANA contains more information than is actually read by the pro gram the following description treats only data that is relevant for DYANA First of all the present version of DYANA does not allow for special endgroups at the N or C terminus of the polypeptide chain Therefore only the entries marked with the keywords ATOMTYPES or RESIDUE are considered The atom types entry starts with a header line with the Fortran format A10 I5 containing the word ATOMTYPES and the number of atom type declarations that will follow The following lines contain atom type declarations in the Fortran format 5X A5 F10 2 215 the atom type the repulsive core radius that will be assigned to atoms of this type a code for hydrogen bond capabilities 1 for hydrogen atoms that can form hy drogen bonds for hydrogen bond acceptors e g oxygens and 0 for at oms that cannot be involved in hydrogen bonds and the order number of the chemical element 0 for pseudo atoms 1 for hydrogen 6 for car bon 7 for nitrogen etc The atom types entry must precede the resid
118. cros can be includ ed in the macro help macro shows all lines of the macro that start with HE condition statement Executes a logical if statement as in FORTRAN 77 1 e the statement is executed if the logical expression condition is true A line with a logical if statement must not end with the word then i 56 if i 1t 0 print i is negative 56 is negative condition then Executes a block if statement as in FORTRAN 77 if mod i 2 eq 1 then print i is an odd number else if def x and exist y then print x is defined and y exists else if s eq then print The variable s is blank end if variable Changes the names of the parameters that are passed to a macro i e the parameters p1 p2 get the names given in the parameter statement The parameter statement must precede all other statements in a macro except var and cannot be used interactively subcommand parameter Performs a plot subcommand Plot commands are described separately in the Graphics section of this chapter 57 INCLAN print quit readline remove return set 58 text level level Writes the text to standard output or into the file with the given filename If the text contains blanks it must be enclosed in double quotes Option ally the importance level of the output can be defined B
119. cs file is removed after the MIF or Postscript file has been produced add replace multiple Creates dihedral angle restraints that include the allowed angle values stored in the standard grid memory A see command grid memory Optionally the new restraints may be added to those already present or the old restraints may be discarded and replaced by those created on the basis of the grid memory If neither the add nor the replace option is set the intersection between the old and new angle restraints will be formed It is possible to generate multiple restraints for one dihedral angle if there are several allowed regions for this dihedral angle present grid correlate grid fragment Es Commands in grid memory By default only one restraint that includes all allowed ranges is created per dihedral angle function A p angle selection all angles in fragment Defines relationships between dihedral angles in a grid search To repre sent a group of dihedral angles as a single degree of freedom in a grid search this command has to be called once for each correlated dihedral angle If exactly one angle within the fragment is selected then it will be related to the parameter p lower case p which will be the single degree of freedom during the grid search and vary from 0 to 27 by the function fp If exactly two angles within the fragment are selected one of which has occurred in a previo
120. d names are initialized when they are first used in grid memory expressions a true initialize grid memory a all angle values allowed a false initialize grid memory a all angle values forbidden a not b not b a b and c intersection of b and c a b or c union of b and c a list contents of grid memory a a remove grid memory a Grid memory expressions must not contain blanks If an angle selection is specified then the operation will be applied to all selected angles By default the operation is performed for all angles in the current fragment The command can be given without any parameters in this case the names of all occupied grid memories are printed tfcut f nax 0 0 test Performs a grid search for the current fragment as defined with the grid fragment command The grid search will be done over all angles in the fragment and with the number of steps given by the variable nstep If the cutoff value for the local fragment based target function fmax 18 positive then all conformations with a local target function value below max Will be considered as allowed Otherwise a conformation will be al lowed if no single restraint violation exceeds the corresponding soft cutoff values defined by the variables soft_upl soft lol etc To avoid excessive computation times for fragments with many angles and or few restraints the calculation is not started if the expected number of grid points to be checked after t
121. d peaks are written file file append Write a chemical shift list traditionally called proton list in XEASY format Eccles et al 1991 Bartels et al 1995 Optionally the output may be appended to an existing file file file o append Writes an upper limit distance constraint file Optionally the output may be appended to an existing file name name ang cor pdb cor 119 Commands A Writes all selected structures to angle ang DG coordinate Cor or PDB coordinate pdb files with names namennn ang na mennn cor or namennn pdb respectively nnn denotes the structure number 120 cut_aco cut_cco cut_lol Variables and Functions DYANA gives the user access to internal variables and functions of the program through system variables and functions With system variables the user can obtain and set parameters of the program with functions the user can obtain the value of parameters of the program but he cannot change them System variables The following is an alphabetical list of all DYANA system variables Cutoff value for angle constraint violations in degrees Only violations larger than this value are listed in the commands structure list and structure violate Initial value 5 0 Cutoff value for coupling constant restraint violations in Hz Only vi olations larger than this value are listed with the commands structure list
122. dinate cor or PDB coordinate pdb files of the structures may be written with file names name ang name cor or name pdb respectively The structures may be analyzed for hydrogen bonds option hbond or for violations of steric lower distance limits option vdw Note Because the target function is re calculated it is important that all constraints used for the calculation of the structures are present and that the same weights are used An overview file may contain four different tables e For each structure the target function value the numbers sums and maxima of constraint violations the output of the structure list command e For each violated constraint the structures in which it is violated by more than the corresponding cutoff value the output of the struc ture violate command By default violations are shown only if they occur in at least one third of the conformers To obtain a listing of all violations larger than the cutoffs the option full must be set e For all pairs of structures the RMSD for the backbone and all heavy atoms output of the rmsd command By default only the average value of all pairwise comparisons is written A table with the indi vidual pairwise RMSD values is created only if the option full is set e For all residues the local RMSD for tri peptide segments and the displacements for backbone and all heavy atoms output of the rmsd command This table is only created if
123. ding average or minimal distances in the selected structures can be created Before calibration the volumes of peaks assigned to pseudo atoms are divided by the number of protons they represent For instance the vol ume of a cross peak between a Leu QQD pseudo atom and a Tyr QB pseudo atom is divided by a6 x 2 12 offset Aw 0 0 Generates constraints for the backbone dihedral angles 6 and y in pro teins by comparing the C chemical shifts with the corresponding ran 85 Commands cluster create 86 dom coil values of Spera amp Bax 1991 Angle constraints are derived according to the rules of Luginb hl er al 1995 The C random coil shifts are relative to internal TSP Optionally an offset Aw is added to the chemical shifts in the proton list A warning is printed for C chem ical shifts that deviate by more than 15 ppm from their random coil val ue file name cluster ps range residue range all residues Calculates the backbone RMSD of the selected structures and performs a cluster analysis on the resulting RMSD matrix The resulting graphics is written into the graphics output file with given file name a GRAF file if the extension is grf a MIF file if the extension is mif or a Post script file otherwise A specific residue range may be specified for the RMSD calculation The y axis of the plot gives the structure numbers and the x axis shows the RMSD with which
124. dyana DYANA version 1 5 sgi double precision Copyright c 1996 98 ETH Zurich dyana gt The title line shows the version number of the program the computer ar chitecture for which it has been compiled and an indication of whether single 32 bit or double 64 bit precision arithmetics will be used dy ana gt is the prompt of DYANA which is shown when the program is ready to accept commands from the user If the prompt does not appear after starting the program the program is not installed correctly Tutorial 10 The command readdata reads the input files for a structure calculation input files are in the example helix directory dyana gt readdata helix Library file soft lib dyana 1 1 1ib dyana lib read 42 residue types Sequence file helix seq read 11 residues Distance constraint file helix upl read 145 upper limits Distance constraint file helix lol read 14 lower limits Angle constraint file helix aco read 26 constraints A command may have parameters that are separated from the command name and from each other by blanks In the above example the readd ata command has one parameter the name of the input data files The readdata command reads the standard residue library dyana lib the sequence of the molecule under study helix seq and files with con formational constraints upper distance limits helix upl lower distance limit
125. e absence of a transposed peak e No possible proton in dimension x No proton chemical shift ex ists within the given tolerance range from the peak position in di mension x In 3D peak lists the hetero atom dimension dimension 3 is coupled to its proton dimension 2 and the message means that the problem occurred in one of both dimensions No comment means that there are more than maxamb assignment possibilities left and that the peak was therefore not considered at this stage At the end of this output the number of peaks belonging to every one of the above categories is given atom glomsa atom list atom mass atom rename Commands atom selection all atoms cutoff c 0 4 threshold 0 4 fraction f 100 Function of the previously separate program GLOMSA Global method for stereospecific assignments G ntert ef al 1991a The selected structures are searched for possible stereo specific assignments of the se lected atoms To be taken into account the difference between two con straints going from a stereo specific atom pair B1 and B2 to another atom a must be at least c A the corresponding average distance difference in the structures must be at least A and the minimal percentage of struc tures in which the sign of the distance difference must be consistent must be larger than f percent atom selection all atoms Lists all selected atoms see chapter Selections This command is use
126. e chemical shifts J Amer Chem Soc 113 5490 5492 Stein E G Rice L M 8 Bringer A T 1997 Torsion angle molecular dy namics as a new efficient tool for NMR structure calculation J Magn Re son 124 154 164 Williamson M P Havel T F amp W thrich K 1985 Solution conformation of proteinase inhibitor Ia from bull seminal plasma by IH nuclear magnetic resonance and distance geometry J Mol Biol 182 295 315 Wiithrich K 1986 NMR of Proteins and Nucleic Acids Wiley New York Wiithrich K Billeter M amp Braun W 1983 Pseudo structures for the 20 common amino acids for use in studies of protein conformations by mea surements of intramolecular proton proton distance constraints with nuclear magnetic resonance J Mol Biol 169 949 961 189 References DA 190 A alias 53 angle fix 76 angle flip 76 angle free 76 angle list 76 angle rename 76 angle set 77 angstat clear 77 angstat list 77 angstat make 77 anneal 77 ask 54 asno 78 atom glomsa 81 atom list 81 atom mass 81 atom rename 81 atom shift 82 atom stereo 83 atom swap 83 atom types 139 atom vdw 83 DS Index Index bmrblist 84 break 54 C calc_all 84 caliba 84 102 calibrate 85 calibrating NOEs 19 cashifts 85 close 63 cluster 86 COFIMA commands string 181 macro 181 ancoma 174 attach 174 bind 174 break 174 change 175 cofima 175 connect 175 constraints 175 coordinates 175 copy 176 1
127. e corresponding numbers are set to 0 and the atom number of the corresponding pseudo atom or 0 if there is no corresponding pseudo atom The nomenclature of atoms in amino acid residues closely follows the IUPAC recommendations The only exception is the backbone amide proton which is called HN instead of H In addition to real atoms the res idue library may contain pseudo atoms identified by the atom type PSEUD that are used in DYANA as dimensionless reference points for distance constraints To avoid nomenclature confusion all atom types and residue entries of the standard residue library file are listed in the following for compact ness two numbers that are not used by DYANA are not printed in the atom lines between the atom type and the x coordinate ATOMTYPES 18 1 PSEUD 10 00 0 0 2 H_AL 1 00 0 1 3 H_AM 0 95 1 a 4 H_ARO 1 00 0 1 5 H_SUL 1 00 0 1 6 H_OXY 1 00 E I 7 CAL 1 40 0 6 8 C_BYL 1 40 0 6 9 C_ARO 1 35 0 6 10 C_VI 1 40 0 6 11 N_AM 1 30 1 7 12 N_AMO 1 30 0 7 13 O_BYL 1 20 l 8 14 O_HYD 1 20 8 15 O_EST 1 20 8 16 S_OXY 1 60 0 16 17 S_RED 1 60 0 16 18 P_AL 1 60 0 15 RESIDUE ALA 4 14 3 13 1 OMEGA 1 2 10 0000 2 1 3 4 0 2 PHI 0 0 0 0000 1 3 5 12 0 3 CHI1 1 3 1 3500 3 5 8 9 11 4 PSI 0 0 0 0000 3 5 12 14 0 le C_BYL 0 6824 1 1357 0 0000 2 3 0 0 0 20 O_BYL 0 1723 2 2550 0 0000 1 0 0 0 0 3N N_AMI 0 0000 0 0000 0 0000 1 4 5 0 0 4 HN H_AMI 0 4226 0 9063 0 0000 3 0 0 0 0 5 CA CALI 1 4530 0 000
128. e modify 90 Modifies distance constraints Redundant and meaningless distance con straints are removed Distance limits with diastereotopic groups are ad justed and or pseudo atoms are inserted if no stereospecific assignment is available Giintert et al 199 1a If the information level is full a detailed listing of all modifications that have been done to upper distance limit constraints such that they allow for both possible stereospecific assignments simultaneously unless a stereospecific assignment is available for a given diastereotopic pair is given For example Modifications for floating stereospecific assignments Atom s A Atom s B Input constraint s modified to A1 Bl Al B2 A2 B1 A2 B2 Ai Bj OA OB Upper HA ASP 1 HD2 3 PRO 2 3 39 3 95 Upper HB2 3 ASP 1 HD2 3 PRO 2 5 50 5 50 3 89 5 41 5 50 4 97 Upper HB2 PRO 2 HG2 3 MET 370300 6 38 Upper HD2 3 PRO 2 QE TYR 19 7 63 8 5 Upper HN MET 3 HB2 3 ET 3 134 95 8 33 3 83 3 45 Upper HB2 3 MET 3 OF ET 3 8 53 6 31 6 53 Upper HB2 3 MET 3 HN THR 4 5 38 5 50 5 50 Upper HB2 3 MET 3 HA THR 4 4 69 5 54 Upper HG2 3 MET 3 HN THR 4 4 14 5 0 Upper HG2 3 MET 3 HB THR 4 5 50 6 38 Upper HG2 3 MET 3 OB ALA 8 6 53 7 40 Upper HG2 3 MET 3 OD TYR 19 7 64 8 52 Upper HG2 3 ET 3 QE TYR 19 7 63 8 5 Upper OF MET 3 HG2 3 ET 16 5 47 6 35 Upper 0G2 THR 4 HE21 2 GLN d 36353 7 39 Upper HN GLU 6 HG2 3 GLU 6 5
129. e only peaks which are different ly assigned in both lists are unassigned range residue range all amino acid residues tfeut fnax 0 0 continue Performs grid searches for all dinucleotide fragments in the given range If the cutoff value for the local fragment based target function max 15 positive then all conformations with a local target function value below fmax Will be considered as allowed Otherwise i e if fmax 0 0 a conformation will be allowed if no single restraint violation exceeds the corresponding cutoff value defined by the variables soft_upl soft lol etc Unless the continue option is set the allowed ranges of dihedral angles will be initialized to allow all possible angle values before the grid searches are started The results include dihedral angle restraints and if possible stereospe cific assignments for the diastereotopic groups in the fragment Checks how well long range distance constraints are supported by other constraints A low score indicates lonely and therefore dangerous constraints with a high impact on the calculated 3D structure distance clear distance compare distance delete distance keep distance list distance correct distance make Es Commands For the distance constraint i going from residue number ri to r with r lt r the score s i is defined as a sum over all other distance con straints j N 1 kiya me 4 1_ yl 2 2 52 a
130. e present in both lists and that differ by more Aw in ppm a warning is printed file file Reads a residue sequence file file unknown erroriwarningiskip error append Reads a upper limit distance constraints file Constraints that involve un known residues or atoms can either cause an error a warning or can be skipped Optionally the distance constraints are appended to those already present file file re unknown erroriwarningiskip error append Reads a file with conformational constraints in XPLOR format Br nger 1992 A simplified version of the atom selection syntax of XPLOR is used Constraints that involve unknown residues or atoms can either cause an error a warning or can be skipped Optionally constraints are appended to those already present read_all readdata redac Es Commands list of files Reads all given angle coordinate or PDB files and stores them into the structure memories File name may contain asterisk and question marks to select several structures at one time e g er ang or er cor The file name extension decides on the format of the individual files files with filename extension ang are read as angle files files with file name extension pdb are read as PDB coordinate files and other files are read as DG coordinate files name Reads input data files with the given
131. e value of the variable with name s or a blank string if no variable with this name exists Character function that returns the value of the variable with name s or s if no variable with the name s exists Integer function that returns the number of seconds since the start of the program Macros Macros are files containing INCLAN statements A macro is called by its 51 INCLAN DI Information level Protocol file 52 name that is identical to its filename except for the extension dya that is required for macro files INCLAN looks for macro files in the directories given by the special variable path or in the explicitly given directory Command line parameters may be passed into a macro Within the mac ro they are available as local variables that are by default called p1 p2 These variable names can be changed with the parameter statement The local variable nparam denotes the number of command line param eters Macros can be called from within other macros On line help in formation may be included into a macro as lines that start with two comment signs Such lines are copied to standard output when one requests help about a macro with the command help macro The special macro init is an initialization macro that is automatically ex ecuted when the program starts Typically this macro sets the system variable path that defines the search path for macro files Standard output This section expl
132. een output full protocol protocol Close protocol file Output from a command is redirected to a given file if the last parameter of the command is gt file Redirect to a new file or overwrite existing file After writing the output the file remains open gt file Redirect to a new file or overwrite existing file After writing the output the file is closed gt gt file Append to an existing file or create new file After writ ing the output the file remains open gt gt file Append to an existing file or create new file After writ ing the output the file is closed Blanks between gt and file are not allowed and that the file name must not end with The file name is optional if it is omitted the output will be redirected to the previously used file When redirection is used all output that would otherwise be sent to the screen is written to the given file Standard output and the protocol file are not used Built in commands The following commands are built into the command interpreter Their names cannot be abbreviated name statement Defines a new alias name i e an abbreviation for the given statement The statement may contain an asterisk to indicate where the com mand line parameters are to be inserted Without parameters alias gives a list of all currently defined aliases alias print S Simulate a pocket calculator S 7 35 53 INCLAN DB ask b
133. elp a question mark may be used 177 COFIMA insert keep link list 178 A Aj A range Insert pseudo atoms This command can only be used with coordinate files The command inserts a new atom with the name A in the centre of the atoms Aj A gt For example the command insert QB HB inserts a pseudo atom QB in the centre of the B protons Aj A range Keep only those atoms distance constraints or angle constraints that match the specification given on the command line As an example keep N CA C deletes all atoms except the backbone atoms N CA and C in amino acid residues When working with distance constraints distance constraints with one or both atom names matching an atom specification on the com mand line are kept b Aj range Ag rangen Generates covalent connectivities between atoms called A in the residue range range and atoms called A in the residue range range if they are less than b A apart The default for bond length is b 2 5 A This com mand can only be used with coordinate files For example the command link SG SG inserts covalent connectivities between atoms called SG which are less than 2 5 A apart from each other and can thus be used to generate the connectivities that correspond to disulphide bridges range Gives a summary listing of the atoms distance constraints or angle con straints in the giv
134. en residue range by default including all residues The number of atoms distance constraints or angle constraints the number of residues and lists of the occuring atom angle and residue names are given pseudo quit read remove rename A COFIMA A Aj A A3 c3 c4 range Modifies distance constraints from real to pseudo atoms Wiithrich et al 1983 This command can only be used with distance constraints files that contain upper distance bounds Distance constraints involving atoms that match one of the atom specifications Aj As are changed in order to refer to the pseudo atom A and the upper distance bound is increased by a correction Usually this correction is given by cq A if the distance constraint is an intraresidual constraint that involves one of the atoms A3 the specific correction given for this atom is used Terminate the program file Read an input file with Cartesian coordinates distance constraints or an gle constraints The program determines automatically which format the input data file has The allowed formats for Cartesian coordinate files are DG The format used by DYANA PDB The format used by the Brookhaven Protein Data Bank Bernstein et al 1977 with some restrictions AMBER The format used by the molecular dynamics program AMBER Singh et al 1986 very similar to PDB Distance constraints and angle constraints are read in the format used by
135. ery possible peak assignment an entry is made Peaks from the three internal NOAH assignment lists are saved see filter Assignments from the ambiguous and unambiguous list also have the distance constraint in A that was derived from the peak volume and the assignment file file o connect all append Writes a coordinate file in DG Distance Geometry format Optionally the output may be appended to an existing file By default the Cartesian coordinates of the current structure are written The covalent connectivities may be included too Optionally the Car tesian coordinates of all selected structures may be written file file append Writes a lower limit distance constraint file Optionally the output may be appended to an existing file write pdb write peaks write prot write upl write_all Es Commands file file o all append Writes a coordinate file in PDB Protein Data Bank Bernstein et al 1977 format Optionally the output may be appended to an existing file By default the Cartesian coordinates of the current structure are written Optionally the Cartesian coordinates of all selected structures may be written file file o selected append Writes a peak list in XEASY format Eccles et al 1991 Bartels et al 1995 Optionally the output may be appended to an existing file By default all peaks are written Optionally only the selecte
136. es since the last output the number of updates of the van der Waals interaction list since the last output and the number of target function evaluations since the last out put For example step time Epot Ekin Etot rmsdev maxdev up 0 0 000 17817 672 5776 000 23593 672 1 a 200 13 778 4367 090 7321 274 11688 363 2 842 18 576 4 204 400 28 471 2896 928 6002 219 8899 147 2 763 16 301 4 206 600 42 374 2464 380 6988 264 9452 645 2 330 13 941 4 200 800 60 234 2496 055 6167 296 8663 351 2 815 15 211 4 200 1000 76 882 1654 211 322 900 6977 11 2 779 15 591 4 200 All energies are measured in target function units Temperatures are measured in target function units per degree of freedom i e per rotatable torsion angle A warning is printed if in a single time step the value of a dihedral angle changed by more than 35 and an error occurs if the change exceeds 90 minimize Es Commands steps N 100 level L taken from variable level flat n 100 angdev Ad 10 0 vdwupdate N jw 100 Performs N conjugate gradient minimization steps including constraints up to minimization level L The flat parameter is used for the flat stop criterion of the conjugate gradient minimizer It is stopped if within n minimization steps the target function cannot be reduced by at least 1 The van der Waals interaction list is updated every Nygy steps or each time a torsion angle has changed its value by more than Ad degrees si
137. es of one or several global variables are given the values of these variables are displayed Waits for t seconds name Defines a new user defined command within a macro i e a macro within a macro User defined commands defined by subroutine statements are called by their name possibly followed by parameters in exactly the same way as macros User defined commands defined by a subroutine statement are local to the current macro or macros called through it Within a macro a user defined command can only be called after it was defined format Analyzes the command line parameters of the current macro This state ment can only be called within a macro Command line parameters that match with one of the format specifications are removed from the list of command line parameters and assigned to a new local variable The possible format items are name type default Declares a named parameter with the given name type and optionally default value If the default value is ab 59 INCLAN 60 name sent the parameter is required and an error will occur if the parameter is not specified in the macro call The optional second sign after the name indicates that a parameter that matches name but does not contain ar an sign is not recognized otherwise with only one Ga sign after name an error occurs in this situation A local variable with the g
138. ession or variable value assigns a value i e a string or the result of an expression to an external non local variable even if a local variable with the same name exists This command can be used to return values from a macro to the calling macro command swap a b Command to swap two variables var x y Declare two local variables x and y x Sexternal Sa Get value of external variable with name a y external b Get value of external variable with name b external a y Assignment to external variable with name a external b x Assignment to external variable with name b end x 10 y 5 print Before swap x x y y Before swap x 10 y 5 swap x y print After swap x x y y After swap x 5 y 10 Returns from a macro to interactive input Given interactively it exits from the program label continues execution of a macro at the first line that begins with the label Jumps into loops do end do or conditionally executed statements if else end if are not allowed and can lead to unpredictable results A label may consist of letters digits and underscore characters _ A label must be followed by a colon go to cont cont print Now at label cont help parameter plot A INCLAN topic Gives on line help for a given topic With no topic given a list of all available help topics is displayed On line help for ma
139. etween the dihedral angle A A7 A3 Ay and the dihedral angle A A2 A3 As is calculated For instance the dihedral angles in a polypeptide can be calculated with the following command angles CA C N CA Calculate dihedral angles Switch to ANCOMA the part of the program for the manipulation of angle constraint files A A A2 Az A4 b 7 0 range Attach atoms to a structure This command can only be used with coor dinate files The atom A is attached to the atom A3 such that the bond length A3 A equals b the bond angle A A3 A equals T and the dihedral angle A A A3 A if A is omitted or the difference between the dihe dral angles A A A3 A and A Ay A3 Ay if Ay is present equals 6 in this case it is not important which atom is specified by A4 Note that b t and O must be given as real numbers with a period to avoid confusion with the following range specification Normally all atoms must be in the same residue This convention can be circumvented by preceding atom names with a tilde In this case at oms are searched through the list of covalent connectivities which allows to use the attach command also if not all atoms lie within one residue attach HB N CA CB OG1 1 09 110 9 123 0 QTHR Attach the B proton HB of threonine if the heavy atom positions are known A rA ra Insert a specific covalent connectivity between the atom A of residue r and the atom A
140. f possible into the first two lists by this command the test assignment list is exclusively fed by the command assign The AAL is cleared at the beginning of this command but peaks already assigned in the current peak list UAL are not altered by this command even if the peak is differently assigned in the assignment file For every peak entry of the assignment file which may contain several assignments per peak the following procedure is made First all assignments with P gt 2 are discarded Assignments from the UAL of the assignment file which pass this test are transferred to the cur rent peak list UAL If the peak in the assignment file was from the AAL or the TAL two cases are distinguished 1 The peak has only one possible assignment and is either transferred to the unambiguous assignment list if P lt l or unassigned 2 The peak has several assignments left If one of them is much better than the rest i e it has P l and the rest has P gt the peak is unam biguously assigned If not all remaining assignment possibilities are stored into the ambiguous assignment list Usually the relation between the thresholds should be gS lt p in which case the following interpretation may help to understand their signifi cance lo I b certainly probably probably certainly correct correct wrong wrong 0 20 50 80 100 violation Flips planes of aromatic rings of PHE and TYR residues and plan
141. ful to test whether a certain atom selection does select the desired atoms value m 1 0 cluster atom selection all atoms Sets if the cluster is not set the mass of all selected atoms to m In this case all inertia tensors are calculated from the masses and positions of their constituting atoms If the cluster option is set the inertia tensors of all rigid units are set as if the rigid units were spheres of radius 5 A with mass JM where M denotes the sum of the atomic masses within the rigid unit Inertia tensors are initialized in this way when the program starts Atomic masses are initialized to unity The mass does only influ ence the MD calculations name atom selection on off clear 81 Commands atom shift 82 Defines an external name for the selected atoms Not more than one atom may be selected per residue External atom names are used in place of the corresponding internal atom name from the residue library when reading input files and writing output files Initially or after the com mand atom rename clear external and internal atom names are iden tical With atom rename off renaming may be turned off temporarily until it is turned on again by atom rename on or by a new external atom name definition atom rename HB1 HB2 QALA atom rename HB2 HB3 QALA These two statements allow reading input files or writing output files in which diastereotopic B protons are called HB1
142. gle parameter even if it contains spaces Variable substitutions in the text still occur A INCLAN text treats text as a single parameter the single quotes remain part of the text Single quotes are used to delimit FORTRAN 77 character string con stants Variable substitutions in the text still occur Text between a comment sign ff and the end of the line is treated as a comment and skipped by the program Commands preceded by are only echoed if the variable echo has the value full has its special meaning only if it occurs as the first character of a command Istring recalls the last interactive command that started with string P has its special meaning only if it occurs as the first character of a com mand A string replacement executes the last interactive command again af ter replacing the first occurrence of string by replacement The third car et is optional unless the replacement string has trailing blanks A has its special meaning only if it occurs as the first character of a command Variables The command line interpreter allows the use of variables in two different ways e Similar to shell variables in the UNIK operating system as variables whose value can be substituted into the command line In this case the value of a variable is a general character string and has no partic ular type e As variables in FORTRAN 77 expressions In this case the value of a variable mu
143. h patterns which are always specified in points These plot pa rameters are changed implicitly by the scale command or if autoscaling is enabled The values of these plot parameters are available in INCLAN as intrinsic functions plotx0 ploty0 plotx1 and ploty1 Initial values X 250 Y 375 X 250 Y 375 73 INCLAN 74 command name Commands There are two kinds of commands in the program DYANA general built in commands of the command line interpreter INCLAN that are not spe cific to the program DYANA see chapter INCLAN and specific DYANA commands This chapter gives an alphabetical list of the DYANA com mands and the standard macros of DYANA Macros can be found in the macro directory They can be used exactly like ordinary commands and are marked with the symbol The names of DYANA specific commands but not of INCLAN commands or macros can be abbreviated as long as there is no ambiguity The syntax parameters and options of a command are given according to the following scheme parameter default value name value default value option1 option2 option3 option4 command name is the name of the command which may consist of more than one word Parameters and options are given in the form of a table in which the left column gives parameters and options and the right column indicates default values for optional parameters or for re quired parameters In the table above
144. he residue range used for the RMSD calculation is 3 37 and the protein name which is used as output file name is er2 The two proton lists used to assign the peaks lists are called h20 prot and d2o prot proton_nam and the disulfide bridges of Er 2 are included as upper addupl and lower addlol limits distance constraint files ss bond upl and ssbond lol Finally the two assigned reference peak lists are given peak_ref and the calibration option is turned on The reference peak lists are only used to give an overview during the NOAH run In principle NOAH can be used for two different tasks 1 Continue to assign a NOESY peak list or 2 Re assign a peak list to check differenc es in the assignment made manually and automatically In case 1 no reference peak lists may be given parameter peak_ref must be delet ed In case 2 an unassigned version of the peak list must be created first e g read prot h2o read peaks er2_h2o peaks assignment delete write peaks er2_h2o_na For 3D lists the procedure is very similar The format of your peak lists should be included into the peak list itself with the line DYANAFOR MAT format see command read peaks or as a separate parameter plformat into the noah call e g plformat hHN hHC NOAH will produce the following important files e er2 cor The coordinate files of the ten best NOAH structures e er2 ovw Overview file of these 10 conformer
145. he IH NMR spectra of ferrichrome peptides I The non amide protons Biopolymers 17 617 636 DeMarco A Llin s M and W thrich K 1978b 1 5 spin spin couplings in alumichrome Biopolymers 17 2727 2742 Eccles C Giintert P Billeter M amp Wiithrich K 1991 Efficient analysis of protein 2D NMR spectra using the software package EASY J Biomol NMR 1 111 130 187 References 188 Ernst R R Bodenhausen G amp Wokaun A 1987 The principles of nuclear magnetic resonance in one and two dimensions Clarendon Oxford G ntert P 1998 Structure calculation of biological macromolecules from NMR data Q Rev Biophys in press G ntert P amp W thrich K 1991 Improved efficiency of protein structure cal culations from NMR data using the program DIANA with redundant dihe dral angle constraints J Biomol NMR 1 447 456 G ntert P Berndt K D amp Wiithrich K 1993 The program ASNO for com puter supported collection of NOE upper distance constraints as input for protein structure determination J Biomol NMR 3 601 606 Giintert P Braun W Billeter M amp Wiithrich K 1989 Automated ste reospecific IH NMR assignments and their impact on the precision of pro tein structure determinations in solution J Amer Chem Soc 111 3997 4004 G ntert P Braun W amp Wiithrich K 1991a Efficient computation of three dimensional protein structures in solution fr
146. he evaluation of restraints that involve a sin gle torsion angle exceeds the value of the variable gridpoints Simi larly a grid search is aborted if the estimated total computation time exceeds gridtime seconds grid swap gridplot habas Es Commands If the number of grid points to be checked after evaluation of the re straints that depend on a single dihedral angle is larger than N ax or if the estimated computation time for the complete grid search exceeds tmax Seconds the calculation will be stopped If the test option is set the grid search will not be started but the expected number of grid points to be checked after evaluation of the restraints that depend on a single di hedral angle will be printed The grid search is restricted to angle values that are allowed according to the standard grid memory A The resulting allowed angle values from the grid search will again be stored in the standard grid memory atom selection all atoms in fragment Swaps the selected diastereotopic partners in distance restraints and sca lar coupling constants in the current fragment It is sufficient to select one atom of a diastereotopic pair to swap both diastereotopic partners file gridplot ps Produces a plot in FrameMaker MIF if the file extension is mif or Postscript format of the allowed dihedral angle values in the standard grid memory range residue range all amino acid re
147. he new en try for example in the following way e Create a sequence file that contains the new residue type preferably in the interior of the chain i e not as the first or last residue e Using this sequence file and the new residue library in the program DYANA create angle and coordinate files for a conformer with ran domized dihedral angles e Start DYANA again with the same sequence and residue library file read the previously produced coordinate file and write again angle and coordinate files without making any minimization e Check whether the angles and coordinates produced by the second run of DYANA coincide closely with those from the first run If this test fails then there is probably a format error in the new library en try or the ordering rules listed above are violated However this test does not detect errors in nomenclature connectivities or pointers to pseudo atoms e Check the coordinates produced by DYANA on a molecular graphics system for example with the program MOLMOL Koradi ef al 1996 DYANA allows for calculations with more than one molecule through the use of special linker residues These invisible linkers consist exclu sively of pseudo atoms i e they can penetrate the real molecules with out causing any steric repulsion and thus allow the program to formally treat a system of several molecules in the same way as a single molecule 27 Tutorial DI Automatic NOESY assig
148. hemical shift list kt prot read 457 chemical shifts In the above example the pseudo atom and the two protons it represents were assigned simultaneously To prevent NOAH and also the user to assign peaks to the pseudo atom the chemical shift of the pseudo atom should be set back to 999 000 in the proton list Second a rough estimate of all missing chemical shifts can be obtained with the command atom shifts missing 29 Tutorial 30 dyana gt atom shifts missing Residue missing shifts ASP i HN THR 4 HG1 SER 11 HB3 HG HIS 14 HD1 THR 15 HG1 MET 16 HG3 TYR 19 HH GLN 21 HG3 TYR 25 HH THR 27 HG1 THR 32 HG1 THR 33 HG1 PRO 40 HG3 91 7 assigned 14 missing chemical shifts The above listing is usual for what we call a nearly completely as signed proton list Most of the missing proton shifts are those of labile sidechain protons which are not always observable The chemical shifts of the proton list can be checked with atom shifts check dyana gt atom shifts check Atom shift limiti limit2 HB2 TYR TT 4 123 4 100 1 620 QE PHE 21 7 566 TSK 5 560 CG LYS 23 29 777 26 440 20 900 CB ALA 39 25 942 24 200 14 500 H GLU 47 5 784 55550 2 840 HB2 PHE 52 1 1 63 3 920 1 400 Atom Residue Shift Median Spread Peaks HN ASN 3 8 671 8 662 0 014 3 HA ILE 29 4 346 4 344 9053 12 HG2 GLU 31 2 326 2 319 0 015 9 HG2 GLN 32 1 049 1 044 0 012 9 HG3 GLU 35 1 829
149. her input is printed in italics Optional pa rameters are given in square brackets and optional parameters that may be repeated zero or more times are given in curly braces In examples input to the program is printed in bold Courier and output from the program DYANA is printed in regular Courier font Comments suggestions and bug reports are welcome Please send them to Peter Giintert Institut fiir Molekularbiologie und Biophysik HPM G22 Eidgen ssische Technische Hochschule CH 8093 Ziirich Switzerland or by electronic mail to dyana mol biol ethz ch Introduction DM Running DYANA Tutorial This chapter explains how to do some of the tasks that are commonly performed with the program DYANA and its interactive command lan guage INCLAN in the course of an NMR structure determination The example input files used for this tutorial are distributed together with the program and can be found in the example subdirectory of the library directory This is a simple example to introduce a new user to the program DYANA It is not CPU intensive and can be executed interactively on any work station Note however that this example does not provide the recom mended strategy for a protein structure calculation See the next section for a complete prototypical example of a realistic NMR structure calcu lation The program DYANA is started by is the UNIX prompt user input is printed in bold
150. hts all selected distance constraints with w file tolerance d 0 5 LO 0 0 L1 50 L2 2 80 Filters an assignment file see command write ass with respect to the selected structures Mumenthaler amp Braun 1995 The percentage Pi of structures in which every assignment is violated is counted Violations smaller than the tolerance distance d in A are not considered LO L1 and L2 correspond to the percentage thresholds mentioned in Mumen thaler amp Braun 1995 This command makes use of three internal peak lists e Unambiguous assignment list UAL All peak assignments which were unambiguous at some stage of the NOAH calculation and which do not violate the structures This list is in fact simply the as signed peak list e Ambiguous assignment list AAL All peaks with more than one assignment which were used in structure calculations and did not flip Es Commands lead to large structural violations Peaks from this list are added to the normal peak list one entry per possible assignment with nega tive peak numbers They are visible with the command peak list but are not written to disk by write peaks Test assignment list TAL All peak assignment which might be possible and are detected by the command assign They were not used in any structure calculation yet The peak assignments from all three lists were stored into an assignment file in a previous NOAH cycle and are redistributed i
151. iable target function method and then analyzed with regard to the distribution of the values of the dihedral angles Redundant dihedral angle constraints are generated for all resi dues with a local target function value below ang_cut in at least nallow structures These constraints are used to re calculate an ensemble of n structures The procedure can be repeated several times At the end the structures are minimized on the highest minimization level against the original angle constraints The different ang_cut values for every REDAC cycle and the number of structures n are given as parameters to the mac ro redac and nallow is a DYANA variable To calculate 50 structures of the protein Er 2 using one REDAC cycle with an ang_cut value of 0 3 you can write redac er2 schedule 0 3 0 0 0 0 structures 50 The two zeros in the ang_cut list stand for the cycle which uses the re dundant angle constraints calculated previously and the cycle where the structures are minimized at the top level In these two cycles no new re dundant angle constraints are generated Several files will be created 39 66 e The angle files er2a ang er2b ang and er2c ang containing all 15 Tutorial DI Running a parallel calculation 16 50 conformers calculated in the cycles a b and c These files can be reloaded into the structure memory at any time with read ang er2a 99 66 e The overview files er2a ovw er2b ovw
152. ich both constraints would contribute the same amount Initial value 1 Tolerance ranges A between peak positions and proton chemical shifts in ppm In automatic NOESY assignment these values are used for at oms that are already used in peak assignments of the current peak list The value of tolerance is a comma separated list of values for the dif ferent spectral dimensions the first and second numbers apply to pro tons the third number to 1 C or N The second number is for protons that are directly bound to the corresponding BC or PN atom the first number for other protons Initial value 0 01 0 01 0 2 ppm Chemical shift tolerance ranges used to check for the existence of tran posed peaks in 3D peak lists given in the same format as for the variable tolerance Initial value 0 05 0 05 0 5 ppm Same as variable tolerance but the A values given here are used in automatic NOESY assignment for all chemical shifts from atoms which are not used in any peak assignment of the current peak list Initial value 0 04 0 04 0 4 ppm Weight value for contributions to the target function from dihedral angle constraints Initial value 5 0 125 Variables and Functions weight_cco weight_lol weight_ori weight_upl weight_vdw 126 Weight value for contributions to the target function from coupling con stant constraints Initial value 0 5 Weight value for contributions to the target function from
153. in general each iteration of a parallel loop must create an output file As an example a parallel version of the macro calculate from a previ ous section without using the calc_all command is calc para calculate a group of structures Handling groups of structures Ds Tutorial Usage calc_para file lt file gt struct lt n gt syntax file struct i 5 readdata file random all struct Create struct random structures do i 1 struct parallel Parallel loop structure copy i 0 Copy structure to active structure anneal Minimize with standard protocol write cor file i 13 3 cor Save structure end do do i 1 struct Serial loop to collect structures read cor file i 13 3 cor Read structure structure copy 0 i Copy to structure end do overview file The call nproc 3 Calc _para helix will then perform the structure calculation in parallel on up to three pro cessors Some DYANA commands for example calc_all are executed implicitly in parallel if the nproc variable is set to a value larger than one There fore without any change already the simple calculate macro of the pre vious section can perform the same parallel computation as the calc_para macro Besides the current structure structure 0 the program DYANA can store a number of other structures structures 1 N The maximal number of structures N that can be stored depends on the size of the structure and is given by the f
154. in structure calcu lation from experimental NMR data for the pheromone Er 2 from Eu plotes raikovi Ottiger et al 1994 note however that the data set used for this example is not the one described in the publication Data are in the directory example er2 The basic input data files are er2 seq amino acid sequence er2 prot chemical shift list er2_h2o peaks H O NOESY peak list er2_d2o peaks D O NOESY peak list er2 cco vicinal scalar coupling constants In addition there are four DYANA macro files init dya CALI BA dya GRIDSEARCH dya and ANNEAL dya to perform the various steps of the structure calculation The initialization macro init is executed each time the program DYANA is started from this directory name er2 protein name used as file name rmsdrange 3 37 default residue range for RMSD dyanalib read library read seq name seq read sequence This macro defines two variables for later use and reads the library and sequence file of the protein Calibration i e conversion from NOESY peak volumes to upper dis tance bounds is performed by the macro CALIBA read seq name seq read sequence and initialize read prot name prot read proton list read peaks name _h2o peaks assigned integrated caliba read and calibrate first peak list read peaks name _d2o peaks assigned integrated caliba read and calibrate second peak list distance unique keep strongest constraint for each distance wri
155. integer closest to r Logical function that returns 1 if a file with name s is currently open or 0 otherwise Real functions that return the coordinates of the two reference points Xo Yo and X Y in the user coordinate system used for graphics see plot parameters XO YO X1 Y1 Real function that returns a pseudo random number pseudo random numbers are uniformly distributed between 0 and 1 rand n rand n n gt real x sign x X2 sin z sinh sqrt z tan z tanh time val s val s s gt walltime A INCLAN Real function that returns a pseudo random number pseudo random numbers are uniformly distributed between 0 and 1 The random number generator is initialized with the seed n Real function that returns a pseudo random number pseudo random numbers are uniformly distributed between 0 and 1 The random number generator is initialized with the seed n and the result is the n th ran dom number generated from this seed Conversion to real type the argument x must be of type integer or com plex for complex x the real part is returned Returns the absolute value of x times the sign of x if x 0 its sign is taken as positive both arguments must have the same type Sine Hyperbolic sine Square root if z is real it must be non negative Tangent Hyperbolic tangent Character function that returns the current time in the form hhimm ss Character function that returns th
156. ints that involve unknown residues or atoms can either cause an error a warning or can be skipped Optionally the orientation constraints are appended to those already present file file structure n 1 unknown erroriwarningiskip error all Reads a coordinates file in PDB format If there is a DYANA header in the file the target function value will be read from the header Optionally only the n th structure may be read from a multi conformer file Other wise all m structures in a multi conformer file are read and stored as structures 1 m The first structure read will also be stored in the default read peaks Es Commands structure memory 0 The presence of unknown residues or atoms can either cause an error a warning or they can be skipped file file weight w 1 0 filter s 55 no filter format string hH reference integrated assigned append Reads a XEASY Bartels ef al 1995 peak list Volumes are scaled with the weight factor w The filter option allows to skip peaks with com ments that match one of the strings 51 5 For 3D NOESY peak lists the format i e the order in which chemical shifts and assignments are given in the peak list may be specified The format string has one character per dimension that identifies the column of 15N or 3C atoms N or C the column of protons bound to DN or BC H and the column of independent protons h If the for
157. ion from constraints that involve a given residue If a given residue and its closest neighbors have a local target function value below 7 then all dihedral angles of this residue are added to the angle statistics thigh 7 jon 8 0 tend 7 4 0 0 steps N 4000 highsteps N cn NIS minsteps n 1000 relax Performs simulated annealing on the current structure with a total of N MD steps starting with Nhigh MD steps at temperature Thign followed by slow cooling during N Nhign MD steps to a final temperature of Teng Finally n steps of conjugate gradient minimization are added The tem perature is measured in target function units per degree of freedom Op tionally more minimization can be performed in order to relax strong overlaps and constraint violations prior to the start of the MD calcula 77 Commands asno 78 tion The relax option can be useful for larger above 200 residues pro teins if otherwise the maximal length of the pair list would be exceeded distance d max 5 5 structure N assignfile file peakfile ile sortdistance color Determines assignment possibilities for NOESY cross peaks on the basis of chemical shift agreement and short corresponding H H distances in a bundle of conformers This command provides the functionality of the former ASNO program G ntert er al 1993 An assignment of a NOE SY cross peak at position j Wo to a proton pair a B with chemical shifts and a is
158. istances to 10 35 10 65 A for A T and 10 6 10 9 A for C G base pairs file ile o structures N 10 maxwidth A0 270 0 redac append Writes an angle constraint file Optionally the output may be appended to an existing file Optionally redundant dihedral angle constraints for the REDAC strategy G ntert lt Wiithrich 1991 may be derived from the current angle sta tistics and included in the output option redac see the commands ang stat make and redac In order to generate redundant dihedral angle constraints for a given residue its local target function value and that of its immediate neighbors must be below the cutoff value given by the variable ang_ cut in at least N structures Redundant dihedral angle con 117 Commands write ang write ass write cor write lol 118 straints with an allowed range wider than Ad degrees are discarded The parameters structures and maxwidth can only be in conjunction with the redac option file file fixed all append Writes an angle file Optionally the output may be appended to an ex isting file By default the values of all rotatable dihedral angles of the current struc ture conformation are written The values of the fixed dihedral angles e g peptide bond angles may be written too Optionally the angles of all selected structures may be written file Write an assignment file that is used by the NOAH command filter For ev
159. iven name is created and ei ther the value specified by the user or in its absence the default value is assigned to it The value must be compat ible with the given type see below In a macro call a named parameter can either be speci fied anywhere in the parameter list in the form name value or as a positional parameter of the form value at the same position in the parameter list as the corresponding format in the syntax statement Only pa rameters that appear before or see below can be specified as positional parameters without giving their name A name may contain an asterisk to indicate how much it can be abbreviated By default all unambiguous abbreviations are allowed If a name starts with an aster isk then the corresponding parameter is a positional pa rameter that cannot be given in the form name value Declares a literal option with the name A local variable with the given name is created If the option name is present in the macro call this variable is set to 1 i e the logical value true otherwise it is set to 0 name name gt kk Declares a set of mutually exclusive literal options with the names name name gt etc Local variables with the given names are created If one of the option names is present in the macro call the corresponding variable is set to 1 i e the logical value true and the other vari ables are set to 0 Allows for
160. k list is much larger than 5 This can be explained by the fact that NOAH tries to find an assignment for all peaks If there are too many noise peaks NOAH may well find a distorted structure which explains another 95 of the peaks i e scores equally well than the real structures in explaining as many peaks as possible see below For this reason NOAH saves all peaks it has found incompatible into the peak lists incomp x peaks These usually small peak lists should be examined carefully with the spectra with XEASY for example Typical ly many of these peaks can be identified as noise peaks and should be eliminated from the peak list because they disturb the NOAH calcula tions For the others the spectroscopist may search for a previously un assigned proton specially if several incompatible peaks lie on the same shift Once an improved peak list and proton list becomes available a new NOAH calculation should be performed until the percentage of in compatible peaks reaches 1 2 The files reliability x contain the reliability distance of every assigned peak All assignment possibilities which have a minimal violation of less than r A in the structure bundle are listed The parameter r can be given to the command reliability and has a default value of 1 0 A The signif 35 Tutorial DI icance of every entry is illustrated in the following figure Number of conformers in which assignment is Possible violated
161. l axes system of the tensor D Initial value 1 0 Hz Rhombic component Dj of the tensor that relates residual dipolar cou plings to the orientation of the corresponding chemical bond See Eq 7 Initial value 0 0 Hz Random number generator seed Initial value 3771 Cutoff for angle restraint violations for allowed conformations in grid searches Initial value 5 0 Cutoff for scalar coupling constant restraint violations for allowed con formations in grid searches Initial value 0 5 Hz Cutoff for lower limit distance restraint violations for allowed conforma tions in grid searches Initial value 0 1 A 123 Variables and Functions soft_upl soft_vdw tf beta tf type 124 Cutoff for upper limit distance restraint violations for allowed conforma tions in grid searches Initial value 0 1 A Cutoff for steric lower limit distance restraint violations for allowed con formations in grid searches Initial value 0 1 A Value of the parameter B if target function type 3 or 4 used see system variable tf_type Initial value 1 0 Type of target function used for distance constraints The same function al form is used for upper limits lower limits and van der Waals lower limits limiting cases term for a violated upper limit d gt b small violation large violation type d b d gt gt b d b and denote the actual distance the upper distance bound and
162. late it The program can detect this only if the constrained distance depends on one or two dihedral angles Many meaningless intraresidual peaks can thus be eliminated duplicate constraint The same constraint occurs more than once in the input for example because transposed peaks were present in the peak list The number of upper distance limits after doing modifications is given at the end of the table depending on the number of stereospecific assign ments modification may increase or decrease the number of constraints factor f Amy Scales the distance bounds of the selected distance restraints by the fac tor f distance constraint selection 91 Commands A distance set distance stat distance unique distance weight filter 92 Selects all distance constraints that match the given distance constraint selection see chapter Selections tolerance A 0 0 Set the distance bounds of the selected distance restraints to the average distance in the selected structures plus A Lists the total number of selected intra residual sequential medium range and long range constraints If the information level is full these numbers are also given for each individual residue Keeps only the most restrictive distance constraint if several constraints exist for the same atom pair This command corresponds to the first part of the distance modify command w Weig
163. le target function in tor sion angle space Braun amp Go 1985 to find three dimensional struc tures that fulfil the conformational restraints The success rate percentage of structures reaching low target function values of this strategy was in particular for larger B sheet proteins often hampered by the fact that the target function has many local minima into which the conjugate gradient minimizer may become trapped because it takes ex clusively downhill steps A decisive improvement of this situation could be expected from other structure calculation algorithms that have the possibility to escape from unfavorable local minima Therefore we created a new NMR structure calculation program DYANA Giintert ef al 1997 that uses simulated annealing combined with molecular dy namics in torsion angle space torsion angle dynamics i e the numer ical solution of the classical mechanical equations of motion Lagrange equations with torsion angles as generalized coordinates The target function takes the role of the potential energy and the system is coupled to a temperature bath which is cooled down slowly from its initial high temperature thereby allowing the system to cross barriers between local minima of the target function Introduction When compared with other structure calculation algorithms that are based on simulated annealing Briinger 1992 the principal difference of torsion angle dynamics is that it works with i
164. lled x y y If a list data block consists of a sin gle column with n numbers this column is called y and an x column with values 1 2 m is added implicitly After reading a block of list data the graphics system is in list mode and various plot subcommands can be applied to vector expressions formed from the column vectors of the list data block These vector expressions are general FORTRAN 77 ex pressions that are evaluated for all vector elements and where the column vectors X Vj Y gt are denoted by X y1 y2 arc caro clip clip close A INCLAN Besides list data a graphics file can contain plot subcommands and comments starting with but not other commands it is not an INCLAN macro The following alphabetical list contains all plot subcommands They are called from INCLAN in the form plot subcommand parameters and in graphics files in the form subcommand parameters Some of the plot subcommands have different parameters in normal and list mode as indicated by normal mode or list mode at the right margin x y a b b ball draws a circle an ellipse or part of a circle or ellipse with the center at x y and half axes a and b If bis omitted a circle with radius a mea sured in the x direction is drawn Optionally only the part of the ellipse starting and ending with phase angles Q and 6 respectively is drawn The phase angle is 0 on the positive
165. llelism It can be exploited almost ideally for parallel computing and renders feasible applications that have so far been unpractical because of their high computational demands DYANA is written in standard FORTRAN 77 and was implemented on a va riety of computers The program is optimized for shared memory multi processor and vector computers Any reports or publications of results obtained with the program DYANA must acknowledge its use by an appropriate citation P Giintert C Mumenthaler and K Wiithrich Torsion angle dynamics for NMR structure calculation with the new program DYANA J Mol Biol 273 283 298 1997 The structure of this manual is as follows The Tutorial gives an intro duction to the program for first users and also explains how the most common tasks are performed with DYANA The interactive command language INCLAN is described in the next chapter Commands gives a complete list and description of all DYANA commands Variables and Functions gives a complete list of all DYANA system variables and func tions Selections describes the syntax used to select atoms angles peaks distance constraints and structures File formats describes the formats of various data files used by DYANA The supporting program COFIMA is described in separate chapters Installation describes the installation of DYANA on Unix systems In this manual names of commands variables etc and literal input is printed in bold Helvetica ot
166. losed by the convex quadrilateral defined by the four points lx Wi list mode draws B zier spline curves through the points of the given vector expres sions x Vj If no vector expressions are specified splines are drawn through the points of all list columns If the x expression is omitted i e if only a single expression y is given the x coordinates are taken from the x column of the list The number of list points must be 3n 1 with integer n See section mark ae Vi draws an errorbar defined by the given x and y coordinates This com mand can also be used in list mode where x y and y are three vector expressions file fit A INCLAN file reads an input graphics file default extension grf containing list data and plot commands and executes the plot commands in the graphics file Graphics files cannot be nested If no output plot file is open when the file command is executed and if the first plot command in the graphics file does not open an output plot file explicitly a new Postscript output plot file with the name file ps is opened implicitly An implicitly opened output plot file will be closed when the end of the graphics file is reached ab 3 Hitec list mode performs a linear least squares fit of the basis functions given by the vec tor expressions f to the data points with x coordinates y coordinates and errors given by
167. lots which show the distribution of dis tance constraints The first plot shows the number of distance constraints plotted against the residue index difference of the corresponding atoms The second plot shows for every residue the number of intra residual white short range vertically hatched medium range horizontally hatched and long range black constraints file intersection notdiff Lists all differences in the assignments between the current peak list and an external peak list file Corresponding peaks must have the same peak numbers in both lists peak 791 20 HB2 9 HN NOAH RelDis 0 0 20 HB2 28 HN File MinVio 0 0 peak 979 7 HA 10 HN NOAH RelDis HZ 8 HA 10 HN File MinVio 0 0 peak 985 13 HA 12 HN NOAH RelDis 0 0 13 HA 15 HN File MinVio 0 5 Number of equal assignments 608 Number of different assignments 3 The first line of each differently assigned peak contains the NOAH as signment and its reliability distance RelDis provided the latter was previously calculated with the command reliability The second line contains the assignment given in the file together with the minimal vio lation MinVio this assignment would have in the selected structures This violation is calculated on the basis of a distance limit of 5 0 plus pseudo atom correction if appropriate and does therefore not consider the peak volume For the interpretation of the RelDis MinVio combina
168. mats E 3N NM 0 0000 0 0000 0 0000 4 EN HAMI 0 4226 0 9063 0 0000 5 CA CAL 4530 0 0000 0 0000 6 HA HAL 7394 0 5422 0 9011 7CB CAL 2 0038 0 6938 1 2474 8 HB2 HAL 6190 1 7132 1 2776 9 HB3 HAL 6190 0 1818 2 1294 10 QB PSEUD 6190 0 9475 1 7035 11 CG C BYL 3 5302 0 7444 1 3384 12 0D1 O BYL 4 0206 1 3033 2 3432 13 0D2 O BYL 4 1722 0 2231 0 4011 14C CBYL 9587 1 4440 0 0000 150 OBYL 1648 2 3835 0 0000 16 N NMM 3 2772 1 5756 0 0000 CYS RESIDUE CYS 5 15 3 14 OMEGA 2 10 0000 2 1 3 2 PHI 0 0 0 0000 1 3 5 3 CHI1 3 1 3500 lt 3 5 7 4 CHI2 3 0 7500 5 7 n 5 PSI O 0 0 0000 3 5 13 C CBYL 0 6824 1 1357 0 0000 20 OB 0 1723 2 2550 0 0000 3N NAMM 0 0000 0 0000 0 0000 4 EN HAMI 0 4226 0 9063 0 0000 5 CA CAL 4530 0 0000 0 0000 6 HA HAL 7661 0 5112 0 9103 7 CB CAL 2 0187 0 7882 1 1831 8 HB2 HAL 6627 1 8178 1 1483 9 HB3 HAL 6627 0 3594 2 1199 10 QB PSEUD 6627 1 0886 1 6341 11 G S RED 3 8479 0 7581 1 1379 12 HG H SUL 4 0261 1 4888 2 2348 13C CBYL 9334 1 4526 0 0000 140 BYL 1232 2 3781 0 0000 15N NMM 3 2494 1 6072 0 0000 CYSS RESIDUE CYSS 4 14 3 13 OMEGA 2 10 0000 2 1 3 2 PHI 0 0 0 0000 1 3 5 3 CHIL 3 3500 3 5 7 4 PSI O 0 0 0000 3 5 22 C CBYL 0 6824 1 1357 0 0000 20 OBYL 0 1723 2 2550 0 0000 3N NAMM 0 0000 0 0000 0 0000 4 EN HAMI 0 4226 0 9063 0 0000 5 CA CAL 1 4530 0 0000 0 0000 6 HA HAL 1 7661 0 5112 0 9103 7 CB CAL 2 0187 0 7882 1 1831 8 HB2 HAL 1 6404 1 8093
169. me 51 INCLAN special characters 1 41 40 41 40 AO 40 40 40 41 40 4l 1 40 gt 41 INCLAN special variables echo 44 erract 45 info 45 nparam 45 nproc 45 pl p2 45 path 46 prompt 46 protocol 46 timing 46 information level 52 init 98 K keep 98 kringle 98 L longrangeplot 99 md 99 minimize 101 N noahmin 103 non standard residues 26 Ds Index nstep 123 O ori_axial 123 ori rhombic 123 output redirection 53 overview 103 P pl p2 45 parallel calculation 16 peak abs 104 peak create 104 peak delete 105 peak deviations 105 peak distance 105 peak list 105 peak scale 105 peak select 105 peak unassign 105 peak unique 105 plot arc 63 caro 63 clip 63 comment 64 cross 64 curve 64 dot 64 errorbar 64 file 65 fit 65 frame 66 function 66 label 66 line 66 mif 67 plus 67 polygon 67 ps 68 195 Index 196 rectangle 68 scale 68 set 69 shape 69 spline 69 square 69 text 70 triangle 70 write 70 plot parameters align 70 angle 71 autoscale 71 border 71 color 71 dash 72 fill 72 font 72 linewidth 72 marksize 72 mode 72 rotate 73 textsize 73 weight 73 KO 73 x0 73 X1 73 xl 73 YO 73 yO 73 Y1 73 yl 73 plots 21 protein structure calculation 11 protocol 52 R ramachandran 106 random_all 106 randomize 106 read aco 106 read ang 107 read cco 107 read cor 107 read lib 108 read lol 108 11
170. meter dt specifies the only initial value of Ar The van der Waals interaction list is updated every N q steps or each time a torsion angle has changed its value by more than Ad degrees since the last update of the an der Waals interaction list The leap frog algorithm is used to perform the torsion angle dynamics steps Usually torsional accelerations are computed on the basis torsion al velocity values that are linearly extrapolated from those half a time step earlier Optionally it is possible to use more exact values which are calculated iteratively Mathiowetz et al 1994 The molecular dynamics simulation starts at time fg with random tor sional velocities chosen as Gaussian random variables such that the ini tial temperature kinetic energy per degree of freedom is Tp unless the continue option is given When a calculation is continued the veloc ities from the end of the previous md command are used and all param eters that are not given explicitly are kept at the values of the previous md command The parameters tinit and estart cannot be used together with the option continue One line of output is written every n time steps giving the current step current time potential energy i e target function value kinetic energy total energy the root mean square torsion angle change per time step in degrees averaged over all time steps since the last output the maximal torsion angle change per time step in degre
171. mmand words are case sensitive Many commands allow for the specification of a residue range denoted 6699 66 39 66 29 by range which consists of one or more of the elements r r ar rir or R separated by at least one blank 12 Residue 12 12 20 25 Residues 12 20 21 22 23 24 25 QTHR All residues with name THR 20 25 QTHR All residues with name THR and numbers 20 25 acY All residues with names that do not start with CY The default residue range that will be used if no residue range is speci fied includes all residues For many commands all selected atoms must be in the same residue This convention can be circumvented by preceding certain atom names with a tilde In this case atoms are searched through the list of covalent connectivities When using covalent connectivities must of course be present either they can be read from a DG coordinate file or they can be generated using the connect bind or link commands The output of those commands that give interesting output can be redi rected to disk files To do this the last parameter on the command line must be gt file here and in the following items given in brackets are optional which writes the output to a new file or gt gt file which ap pends the output to an existing file Note that no space is allowed be tween the gt sign and the output file specification If the output file
172. n the current set or in the new set _ atoms in the current set but not in the new set atoms in the current set and in the new set Operators are always evaluated from left to right The current atom set is the set of atoms defined by what precedes the operator The new atom set is the set of atoms defined by what follows the operator An empty atom selection selects all atoms HA atoms called HA HA HB all atoms called HA or HB HA ALA 10 20 HA in ALA of residues 10 20 HA ALA 10 20 HA in ALA except in residues 10 20 N CA C 15 17 Hk Q all backbone atoms and the sidechain heavy at oms of residues 15 and 17 The command atom list can be used to check atom selections Angle selections follow the same syntax as atom selections except that angle names instead of atom names are specified The command angle list can be used to check whether angle selections Peak selections are made with the peak select command and consist of two atom selections that are separated by a comma The comma may be omitted if both atom selections consist of a single atom name In addi tion peak selections may contain one or several of the following condi tions levels m n Select only peaks between residues that are between m and n residues apart volume V nin Vmax Select only peaks with volume between V nin and Vmax fraction p Distance constraint selection Structure selection Ds Selections Select randomly only the fraction
173. name If name has an extension i e if it contains a a file with the corresponding format as given by the extension is read Otherwise the sequence file name seq and if avail able the upper limits distance constraints file name upl the lower lim its distance constraints file name lol and the angle constraints file name aco are read If no residue library is present the standard DYANA library dyana lib is read in advance name name schedule schedule 0 4 0 0 0 0 structures n 50 steps N N gt N3 150 400 800 minimizer macro vtfmin Performs REDAC cycles Giintert amp Wiithrich 1991 with n structures according to the given schedule Overview and angle files of every cycle are written to the files name ovw and name ang where the asterisk is replaced by a b c etc for successive cycles The schedule is a comma separated list of ang_cut values that will be used to generate redundant dihedral angle constraints Structures are calculated using the given macro for minimization This macro must accept the same param eters as the standard variable target function minimization macro Vt fmin A zero or negative ang_cut value means that no redundant angle constraints will be generated in this cycle The next cycle will therefore use the original angle constraints to minimize the current structures on the last level during N3 iterations Otherwise i e if ang
174. nce the last update of the an der Waals interaction list If the information level is normal or higher one line of information will be printed out as in the following example from the macro vtfmin Minimization standard strategy lev upper lower vdw angle target funct grad up f stop act act act act begin end end 0 115 13 0 0 313 26 84 9 282 65 0 13 1 8E 2 0 150 maxit 1 271 47 0 0 925 73 84 5163 09 3 79 0 16 34 150 maxit 2 299 51 0 O 1067 81 84 5 24 71 3 79 0 11 18 150 maxit 3 381 57 0 0 1240 92 84 8 694 27 4 22 0 27 20 116 flat 4 431 74 0 0 1335 90 84 10 21 83 4 47 0 16 9 130 flat The first column gives the minimization level Then there are four times two columns containing each time the total number of constraints and the number of active constraints for the upper limit constraints the lower limit constraints the intrinsic van der Waals lower limit con straints and the angle constraints Active constraints are those that yield non vanishing but often small contributions to the target function Following this data the value of the target function at the beginning and at the end of the minimization step is given accompanied by the norm of the gradient of the target function at the end of the minimization step the number of updates of the van der Waals contact list the number of target function evaluations and a stop criterion code The following stop crite ria codes may occur gradtl The squa
175. nment NOAH 28 Each of the linker residue types in the standard library has one rotatable bond The residues are lt PL PLM to link an amino acid residue to a generic linker e NL NLM to link a nucleotide residue to a generic linker LL LL2 and LL5 LLM LLM2 and LLM5 generic linker residues with 1 2 and 5 A bond lengths respectively and 90 bond angles lt LP LPM to link a generic linker to a following amino acid residue e LN LNM to link a generic linker to a following nucleotide residue There are two forms of each linker residue type The normal form is used for minimization and in TAD calculations with spherical inertia tensors the default In TAD calculations with inertia tensors directly derived from the atomic masses and positions the forms given in parentheses must be used A sufficient number of these linker residues must be used between two molecules such that no artificial constraint on the relative positioning of the two molecules with respect to each other is introduced by the finite length and flexibility of the stretch of linker residues To treat for example a system consisting of a double stranded DNA of residues 1 14 and 101 114 and a protein starting with residue 200 the sequence file could look like this GUA 1 ADE ADE ADE GUA CYT CYT ADE THY THY ADE GUA ADE GUA 1st DNA strand NL 50 Tali Tali
176. none of the above end if 25 Tutorial DA Creating non standard residues 26 Repeated execution of commands is achieved by forming loops with the do statement Loops executed a predefined number of times have an in teger loop variable do i 1 20 print i i end do Here the loop variable i runs from 1 to 20 in steps of 1 A loop that is executed until a termination condition is met can be con structed as follows do if x gt 100 0 or finished break end do The break statement exits from a loop Unconditional jumps are possible by virtue of the go to statement do i 1n if err go to cleanup end do cleanup print Error in the loop The go to statement transfers the program flow to the position indicated by the label cleanup Non standard residue types can be added to the residue library as addi tional entries The procedure to add a new residue type to the library is as follows see section File formats for a description of the format of the residue library file Create Cartesian coordinates for all atoms of the residue for example with a molecular graphics program or using the attach and insert com mands of the program COFIMA Bond lengths bond angles and chirali ties of this structure must be correct but the conformation i e the values of the dihedral angles does not matter The coordinates of the overlap at oms at the beginning and at the end of the residue for example N C
177. nternal rather than with Cartesian coordinates The covalent structure parameters bond lengths bond angles chiralities and planarities are always kept fixed at their op timal values The strong potentials required in conventional Cartesian space molecular dynamics to retain the covalent structure and concom itantly the high frequency motions caused by them are absent in torsion angle dynamics This results in a simpler potential energy function and in longer permissible time steps for the numerical integration of the equations of motion and thus in a much higher efficiency of the algo rithm To fully exploit the great potential advantages of torsion angle dynamics a careful consideration of its implementation was required because the Lagrange equations of motion with torsion angles as degrees of freedom are much more complex than Newton s equations in Cartesian coordi nates A naive implementation of torsion angle dynamics Mazur et al 1991 would entail in every time step the solution of a system of N linear equations N being the number of degrees of freedom and thus require a prohibitive computational effort proportional to N In contrast DYANA uses a fast recursive implementation of the equations of mo tion originally developed for spacecraft dynamics and robotics Jain et al 1993 with a computational effort proportional to N In addition the new structure calculation program DYANA incorporates a method for the
178. nts as conditions of if statements as command parame ters when a numeric value is expected and as substring and element in dex expressions An expression is built according to the rules of FORTRAN 77 from con stants variables and function calls These basic items can be combined by operators gt ET Sf pairs eq ne MW le ge gt and or not eqv neqv er lt gt gt and grouped by parentheses There are the following differences to the rules of FORTRAN 77 e The data type double precision is not supported e The data type logical is represented by the integer values O false and 1 true Any integer expression can be used in place of a logical expression with 0 representing false and all other values repre senting true e Variable function and operator names are case sensitive The names of logical operators and intrinsic functions must be written in lower case e The logical operators I lt lt gt gt 8 and can be used in place of its respective FORTRAN 77 equiva abs x acos r aimag c aint anint r asin atan A INCLAN lents eq ne te le ge gt and or and not e All FORTRAN 77 intrinsic functions except dble dp
179. o OO 0 11 0D PSEUD 3 5334 0 8057 1 5014 0 0 0 0 13 12 0E PSEUD 6 0510 0 9730 1 8132 0 0 0 0 B 13 OR PSEUD 4 7922 0 8893 1 6572 0 0 0 0 0 14 CG C VIN 3 3898 0 7961 1 4835 7 15 22 0 0 15 CD ARO 3 9068 1 4012 2 6111 14 16 17 0 0 16 HD1 HARO 3 2356 1 8074 3 3679 15 0 0 0 al 17 CEL CARO 5 3328 1 4960 2 7877 15 18 19 0 0 149 File Formats VAL ADE 150 28 GUMUN N DODIDUBPWNPRPOWOWM NDOT BWDHY 21 22 RESI N DCODADUNPWNPRPDOODINDUTUAPWNHPFPODINDADOBWNE NNN wNH HE1 H_ARO CZ C_VIN CE2 C_ARO HE2 H_ARO CD2 C_ARO HD2 H_ARO OH O_HYD HH H_OXY N N_AM am un H Q w EE bo KK cit OOTAaATAaATZO DODANAUWASEZ PERANG wNH qm A uva Enmzo no I I kh ll PEE Pp Pp pp SS See Q a N Q I Q N w T Q O Q U U a N N on I a Il u u to po p ss EE E DUE ADE ZETA ALPHA BETA GAMMA DELTA NU2 NU1 CHI EPSI C3 03 P OP1 OP2 05 Co H5 H5 Qs ca H4 ES H3 c2 H2 H2 Q2 ci Hl 04 ooooooo00 3 PERS Pee Bes o Mc py EEE Qvmo un gt E 5 Y ma Ke waal lolol sl pp PP Pp Q D E Er E OECS CIEVET E E EOE EV E TIRE oO Cr AD T C4 WENN NW HB 0 01 01 Y CIA ERA AS I ALO FA eo AO E NR Oooo DD OD Om Oxo Om N WN 7543 1417 6654 3477 2394 8311 4885 9579 0013 2356 3231 ooo c
180. olor 5 An assignment was found neither by the user nor by as no The color codes 1 3 are applied only to peaks of the input peak list All additional asno assignment possibilities get the color code 6 except those which are already assigned by the user dist d 5 0 transposed Ar 10000 oneass Finds new possible assignments using the actual peak and proton list and stores them into a test assignment list see command filter for informa tion on different internal peak lists used by NOAH For all unassigned peaks a list of possible proton pairs that have chemical shifts within A from the peak position is made The value for A is taken from the vari able tolerance if at least one peak in the input peak list was assigned to the corresponding proton and from the variable tol_una if the proton was never assigned to any peak in the input peak list This allows to dif ferentiate between proton shifts whose position is precisely determined in the spectrum and those which were determined in another spectrum and may be shifted in the actual spectrum The selected structures are used to reduce the list of possible assign ments in the following way For each proton pair the corresponding up per distance limit Obsdis d seua Where dpseug is the pseudo atom correction if appropriate is determined and a tolerance distance dig is added Mumenthaler amp Braun 1995 If none of the structures can fulfil this enlarged distance limit
181. om mand line that matches the atom name Covalent connectivities are only generated between atoms that are in the same or in sequentially neigh boring residues To generate other connectivities the commands bind and link can be used Aj A range List the distance or angle constraints involving the specified atoms or an gles This command can only be used for distance constraint or angle constraint files Aj Ag range List the atom names residue names and numbers Cartesian coordinates and if present covalent connectivities of the specified atoms This com 175 COFIMA E copy delete difima directory disconnect 176 mand can only be used with coordinate files A r Ag r A3 Copy the atom A of residue rj i e its Cartesian coordinates to atom A of residue r This command can only be used with coordinate files It adds a new atom to residue rp If A is omitted the name of the new atom will be A If Az is given the new atom will be inserted after Az in r3 otherwise as the last atom of the residue Aj A range Delete the specified atoms or constraints When working with Cartesian coordinate files all atoms whose name matches one of the atom specifi cations on the command line are deleted When working with distance constraints all distance constraints for which one or both atom names match an atom specification on the command line a
182. om 131 rad 131 rmsd_bb 131 rmsd_bbdev 131 rmsd_hv 131 rmsd_hvdev 131 mam 131 mum 131 seldis 132 selected 132 shift 132 stereopartner 132 Ds Index tf 132 tfcalc 132 tfmin 132 tfres 132 timestep 132 tolcco 132 DYANA system variables ang_cut 122 cut_aco 121 cut_cco 121 cut_lol 121 cut_ori 122 cut_upl 122 cut_vdw 122 hb_ang 122 hb_len 122 level 122 maxamb 123 obsdis 123 seed 123 soft_aco 123 soft_cco 123 soft_lol 123 soft_upl 124 soft_vdw 124 tf_beta 124 tf type 124 tol_transp 125 tol_una 125 tolerance 125 F filter 92 flip 93 forall 94 G graf 94 grid correlate 95 grid fragment 94 95 193 Index 194 grid memory 96 grid search 96 grid swap 97 gridpoints 122 gridtime 122 groups of structures 17 H habas 97 hbond 98 117 if command 57 INCLAN control statements 25 macro 13 variables 24 INCLAN intrinsic functions abs 47 acos 47 aimag 47 aint 47 anint 47 asin 47 atan 47 atan2 48 char 48 cmplx 48 conjg 48 cos 48 cosh 48 cputime 48 date 48 def 48 dim 48 exist 48 existfile 48 exp 48 external 48 fitchisq 48 fiterr 49 fitpar 49 fitprob 49 getenv 49 getpid 49 global 49 ichar 49 if 49 index 49 indexr 49 int 49 len 49 length 50 lenstr 50 log 50 log10 50 macro 50 match 50 max 50 min 50 mod 50 mtime 50 nint 50 opened 50 plotx0 50 plotx1 50 ploty0 50 plotyl 50 rand 50 real 51 sign 51 sin 51 sinh 51 sqrt 51 tan 51 tanh 51 time 51 val 51 wallti
183. om nuclear magnetic resonance data using the program DIANA and the supporting programs CALIBA HA BAS and GLOMSA J Mol Biol 217 517 530 G ntert P Qian Y Q Otting G M ller M Gehring W J amp W thrich K 1991b Structure determination of the Antp C39 S homeodomain from nuclear magnetic resonance data in solution using a novel strategy for the structure calculation with the programs DIANA CALIBA HABAS and GLOMSA J Mol Biol 217 531 540 G ntert P Mumenthaler C amp Wiithrich K 1997 Torsion angle dynamics for NMR structure calculation with the new program DYANA J Mol Biol 273 283 298 G ntert P Billeter M Ohlenschl ger O Brown L R amp Wiithrich K 1998 Conformational analysisof protein and nucleic acid fragments with the new grid search algorithm FOUND J Biomol NMR in press Jain A Vaidehi N amp Rodriguez G 1993 A fast recursive algorithm for mo lecular dynamics simulation J Comp Phys 106 258 268 Kie ling I amp Lowes M 1987 Programmierung mit FORTRAN 77 Teubner Stuttgart Koradi R Billeter M amp Wiithrich K 1996 MOLMOL A program for dis play and analysis of macromolecular structures J Mol Graph 14 51 55 Laskowski R A MacArthur M W Moss D S amp Thornton J M 1993 PROCHECK A program to check the stereochemical quality of protein structures J Appl Cryst 26 283 291 Lugin hl P Szyperski T amp W thrich
184. on both axes default If the option grid is present a fine grid is drawn If the option zero is present fine lines will be drawn along x O and y 0 if they fall within the frame ae plots the functions given by the expressions f x axis position text labels the given axis by placing a tic and the fext at the given position The parameter axis can have the following values x or bottom Label the x axis i e the horizontal line at y position Yo y or left Label the y axis i e the vertical line at x position A top Label the horizontal line at y position Y right Label the vertical line at x position X If text is blank only a tic is set KIYI Koyo normal mode line mark mark mif plus polygon A INCLAN draws a line that connects the points x4 Y1 X y by straight line segments lx y list mode draws straight lines through the points of the given vector expressions x Y If no vector expressions are specified straight lines are drawn through the points of all list columns If the x expression is omitted i e if only a single expression y4 is given the x coordinates are taken from the x column of the list xy normal mode where mark stands for either dot square caro plus cross or trian gle marks the position x y with the corresponding symbol The size of the symbol is determined by the
185. on the distances involving pseudo atoms located centrally with respect to the diastereotopic substituents No distance limit is indi distance scale distance select Es Commands cated if none will be imposed because the modified distance limit s would be meaningless In addition to the modifications done to account for the absence of ste reospecific assignments the command distance modify detects and removes meaningless constraints in the input A table is given if the in formation level is full For example Meaningless distance constraints limit diagnosis Upper HA ASP 1 HB3 ASP AN 3 21 duplicate constraint Upper HA ASP 1 HB3 ASP 1 3 21 no restriction Upper HA ASP 1 HD2 PRO 2 4 20 duplicate constraint Upper HA ASP 1 HD2 PRO 2 4 20 no restriction Upper HA ASP Ji e DS PRO 2 3 21 duplicate constraint Upper HB2 ASP 1 HB3 ASP 1 2 40 duplicate constraint Upper HB2 ASP 1 HB3 ASP ih 2 40 fixed distance Upper HB2 ASP 1 HD2 PRO 2 5 50 duplicate constraint umber of modified constraints 597 Distance constraints can be meaningless for one of the following rea sons fixed distance The constraint concerns an interatomic distance that can not be varied by changing the rotatable torsion angles Examples of this sort are constraints between geminal hydrogen atoms or constraints between atoms of the same aromatic ring no restriction The constraint is such that there exists no conformation that would vio
186. ons character strings must be enclosed in single quotes dyana gt s Dyana dyana gt l lenstr s ERROR Illegal expression lenstr s is an error because the variable s does not contain a quoted string lenstr is an INCLAN function that returns the length of a string i e the index of its last non blank character The correct use of simple unquoted strings in an expression is dyana gt l lenstr s dyana gt show 1 1 5 Single quotes do not inhibit variable substitutions INCLAN provides a full set of control statements to direct the program flow These are used mainly in macros i e in collections of INCLAN statements that form new commands which can be used in the same way as basic commands Since control statements are not used interactively the program prompt dyana gt will no longer be shown Commands can be executed conditionally by virtue of the if statement which has the same form as in FORTRAN 77 if i gt 20 print i is larger than 20 if i 1t 0 then print i is negative else if i 1t 10 and mod i 2 eq 0 then print i is less than 10 and even else print i is none of the above end if Alternatively comparison and logical operators can also be given in the form of the C programming language if i gt 20 print i is larger than 20 if i lt 0 then print i is negative else if i lt 10 amp amp mod i 2 0 then print i is less than 10 and even else print i is
187. oo ooo amp 9681 6348 4817 7976 0167 5255 4216 4568 2157 8362 2779 5099 1963 1027 3731 4019 7912 5965 0000 0659 0000 1209 3784 Lo 0 0 02 0 19 0 42 39 52 0000 0000 3500 3500 3500 0000 ka 00 90 00 O 0 0 0 2 0 18 i 2 2 2 44 38 lt 57 0000 0000 0000 0000 0000 0000 0000 0000 0000 oo NN 97 97 37 28 06 65 13 25 48 8l 33 83 57 83 41 27 23 19 73 20 70 3 90 67 53 60 85 84 08 wWwuuwHrNH 57 50 00 63 00 70 67 67 54 73 18 9 89 6 94 80 26 13 64 40 35 56 aa 0 0471 0 3652 ila a1 0000 0000 0000 N OO 00001 1G ON ooo o000o0rooorooooo0rNrrrunnR HAHAHA NVWOUWRIDAWH 6734 8244 7019 5253 9912 2132 iA 15 20 0000 0000 0000 0000 0000 9180 1833 0997 2105 1531 2052 0561 2932 2821 1589 0107 2341 2144 1818 0000 0000 0000 IT 13 37 19 21 23 HN N NORPOFPFOUUMOWRRPTONANDWWWDY N 0 20 21 20 23 27 H N p DORPVDOVDWOHDDOCONDVVODOWOHDOLOWO hop N E oOOoosowWoNooomS JOoo gt woo NNER BW WO 0 24 22 23 oo wooo0o0 m m m N OONOOOONOOOWOOOHONSOUOO Js m N N DONGO GOOD DO GOWO GO DINGGO Ww Wu N H Oo0O0o0o0000000BRO0O000O0O0O000O0O H H w oooouwooowo Jow
188. oooHhHooonoo ooooooo00o0000 j H oOoOOOOoOHOoOoNOOonm HH 00w0wO0wwOooooooo RRR ooooooo HH HH oooooovvooooocoooooooooooo Ms File Formats 24 N3 NA 2 8170 2 0867 0 9370 23 235 0 0 0 25 C2 CARO 4 1036 2 4736 1 0196 24 26 27 0 0 26 H2 HARO 4 6227 2 4240 1 9654 25 0 0 0 0 27 N1 NA 4 9152 2 9484 0 0482 25 28 0 0 0 28 c6 CARO 4 4155 3 0749 12114 27 29 33 0 0 29 N6 NA 5 2378 3 5526 2 1808 28 30 31 0 0 30 H61 HA 6 1905 3 8024 9570 29 0 0 0 32 31 H62 HA 4 8967 3 6581 3 1256 29 0 0 0 32 32 06 PSEUD 5 5436 3 7303 2 5413 0 0 0 0 0 33 C5 CARO 3 0716 2 6978 4177 23 28 34 0 0 34 N7 NA 2 2897 2 7047 2 5414 33 35 0 0 0 35 c8 CARO 1 1186 2 2402 2 1184 22 34 36 0 0 36 H8 HARO 0 2357 2 0985 2 7240 35 0 0 0 0 37 03 OEST 2 6047 0 9094 1 3466 13 38 0 0 0 38 P PAL 3 5778 2 1447 1 6417 37 0 0 0 0 RADE RESIDUE RADE 10 38 3 37 1 ZETA 0 0 0 00 wo 9s 3 e 0 2 ALPHA 0 0 0 00 gt Ba e To 3 BETA 0 0 0 00 A 4a O 4 GAMMA 0 0 0 00 e E A 0 5 DELTA 0 0 0 00 7 11 a 6 NU2 O 0 0 0 11 13 15 19 36 7 HOXI 0 0 0 00 13 dee I7 18 38 8 NUL 0 0 0 0 13 is 19 t 36 9 CHI 0 0 0 00 21 19 22 23 36 10 EPSI 0 0 0 00 it 43 37 36 1 C3 CALI 0 9681 5 8551 2 5577 2 0 0 0 2 03 OEST 0 6348 4 7127 1 7719 1 3 0 0 0 3P PAL 0 4817 3 6875 2 2842 2 4 5 6 0 4 OP1 O BYL 7976 4 3635 2 3259 3 0 0 0 0 5 0P2 O BYL 0 0167 3 0215 3 5214 3 0 0 0 0
189. ore a reliability distance of 100 Peak 816 was not assigned by 36 Identifying dangerous NOAH assignments Well supported long range NOEs Single long range NOE that is not supported by other NOEs to adjacent residues Ds Tutorial the spectroscopist the r is missing probably because it is an overlap of the two possible NOEs Peaks 817 and 824 have high reliability dis tances and should be safe Experience has shown that the reliability distance is quite efficient in identifying uncertain assignments Over 75 of the peaks that were dif ferently assigned by NOAH than by the spectroscopist had a reliability distance of O A in a recent study Mumenthaler et al 1997 Therefore you may want to use the command keep after the command reliabili ty which will delete all assignments with a reliability distance equal or below a user given threshold When analyzing the NOAH output one must keep in mind that the elimi nation of erroneously assigned constraints through contradiction with correct constraints will in general be less efficient in regions of low NOE density such as chain ends surface loops or the periphery of long side chains than in the well defined protein core The final distance constraint list should therefore be checked by the com mand distance check that calculates a score for every long range dis tance constraint High scores indicate that there are many other long range distance constraint
190. oth as signments Wiithrich er al 1983 Giintert et al 1991a By default the distance modify command assumes that none of the diastereotopic partners are stereospecificly assigned Therefore all ste Calibrating NOEs Ds Tutorial reospecifically assigned atom pairs should be declared before distance modify is called This is done with the atom stereo command e g atoms stereo HA1 22 30 38 atoms stereo HB2 2 5 6 7 14 16 19 20 24 25 35 37 atoms stereo HD2 35 40 atoms stereo OD1 24 A list of all diastereotopic partners with and without stereospecific as signment can be obtained with atoms stereo list Calibration i e the conversion of peak intensities into distance con straints has become very versatile in DYANA Peaks from the peak list are selected with any criterion command peak select and then cali brated with any monotonically decreasing function command cali brate You can therefore define your own calibration classes and calibration functions A macro Caliba performs a standard calibration of the current peaks us ing three different calibration classes One for NOEs assigned to back bone protons one for NOEs assigned to the more flexible side chain protons and one for NOEs assigned to methyl groups The calibration functions used for these three classes are V A r V Bir V C r where V is the peak volume and r is the corresponding distance The pa rameters A B and C are either given by the user o
191. owoooHrooonoo N w ooooooooNDooovcooWoHhoooHrooonoo H BR BR Ww OOOO OOO OOSC OOD OOO OCOD ODO OOO OOO O00 000000 HH Sooo oc eo oe oO eC II ID Ms File Formats 28 H41 H_AMI 5 1756 3 5735 2 3597 27 0 0 0 30 29 H42 HAMI 4 2049 3 5955 3 8149 27 0 0 0 30 30 Q4 PSEUD 4 6903 3 5845 3 0873 0 0 0 0 0 31 C5 CARO 2 0221 2 6890 2 8462 26 32 33 0 0 32 H5 HARO 1 9484 2 9039 3 9020 31 0 0 0 0 33 C6 CARO 0 9996 2 2045 2 1264 22 31 34 0 0 34 H6 HARO 0 0488 2 0117 2 6008 3 0 0 0 0 35 03 OEST 2 6047 0 9094 1 3466 13 36 0 0 0 36 P PALI 3 5778 2 1447 1 6417 35 0 0 GUA RESIDUE GUA 9 39 3 38 1 ZETA o Oy gt 1 2 3 6 0 2 ALPHA 0 0 0 0000 2 3 6 7 0 3 BETA O 0 0000 3 6 7 1 0 4 GAMMA 0 0 0 0000 6 7 11 13 0 5 DELTA 0 0 0 0000 7 11 13 38 0 6 NU2 O 0 0 0000 11 13 15 19 37 7 NUL O 0 0 0000 13 15 19 21 37 8 CHI O 0 0000 21 19 22 23 37 9 EPSI 0 0 0 0000 11 13 38 39 0 103 CALI 0 9681 5 8551 2 5577 2 0 0 0 0 203 OEST 0 6348 4 7127 1 7719 1 3 0 0 0 3P PAL 0 4817 3 6875 2 2842 2 4 5 6 0 4 OP1 O BYL 1 7976 4 3635 2 3259 3 0 0 0 0 5 0P2 O BYL 0 0167 3 0215 3 5214 3 0 0 0 0 605 OEST 0 5255 2 5916 1 1193 3 7 0 0 0 7 05 CAL 4216 14869 1 2135 ge Bs at 8 H5 HAL 2 4568 1 8260 1 2707 7 0 0 0 10 9 H5 HAL 2157 0 8907 2 1041 7 0 0 0 10 0 05 PSEUD 1 8362 1 3584 1 6874 0 0 0 0 0 1 047 CAL 2779 0 5959 0 0000 7 12 21 13 0 2 Ha H
192. p of all peaks that would normally be selected This option is useful to sim ulate peak lists variable Select only peaks that correspond to a variable distance peaklist filename Select only peaks that were read from the peak list with the given filename without extension The current peak selection may be combined with the previously made peak selection using one of the operators union Select peaks that are selected by any of the two selec tions intersection Select peaks that are selected by both selections xor Select peaks that are selected in exactly one of the two se lections By default previously selected peaks are not considered The command peak list can be used to check peak selections Distance constraint selections are made with the distance select com mand and follow the same syntax as peak selections except that the con ditions volume V in Vmax and peaklist filename cannot be used The command distance list can be used to check distance constraint selections A structure selection consists of one or several of the following elements separated by blanks m a structure number m n a range of structure numbers M from the structure with number m onwards N from the first up to the structure n An empty structure selection selects all structures 135 Selections A 136 File Formats This chapter describes the format of the input files to DYANA Most input With the exception of residue library angle
193. peak positions and proton chemical shifts after the command assign NOAH variable Number of peaks in the unambiguous assignment list that have a different assignment as in the reference peak list after the command filter Number of peaks Number of peak lists Number of residues Number of selected distance constraints Number of selected structures Number of atoms that are associated with the pseudoato n Number of upper limit distance constraints Numerical constant 1 3 14159 Number of the pseudoatom associated with atom n If no pseudoatom is associated with atom n the function returns 0 Numerical constant 180 n 57 2958 Average of the pair wise backbone RMSD calculated with the command rmsd Standard deviation of the pair wise backbone RMSD calculated with the command rmsd Average of the pair wise heavy atom RMSD calculated with the com mand rmsd Standard deviation of the pair wise heavy atom RMSD calculated with the command rmsd Character function that returns the name of the residue n External residue number of the residue n 131 Variables and Functions seldis selected n shift n stereopartner n tf tf n timestep tfcalc tfmin tfmax tfres n tolcco n 132 Average distance bound of all selected distance constraints Logical function that returns 1 structure n is selected or 0 otherwise Real function that returns the chemical shift of atom n or 999
194. possible if the condition an 2 B 2 0 0 ak AP 3 AO AO is fulfilled Aw and AQ are the first and second component of the sys tem variable tolerance and if the distance between the two protons is shorter than dmax in at least N conformers Giintert er al 1993 Three dimensional spectra are treated analogously The option assignfile generates a file containing all assignments which are allowed by asno that can be displayed in the assignment window of the program XEASY Bartels et al 1995 Additionally a new peak list for XEASY containing the old peak list and all assignment possibilities found by asno is produced by using the option peakfile An assign ment possibility to a proton pair a B leads to a new peak at position 0 mb Assignment possibilities for individual peaks are by default sorted ac cording to the chemical shift deviations or if the option sortdistance 1s set by H H distance values If the option color is set the peaks of the input peak list get a XEASY color code according to the following criteria color 1 The assignment given by the user is found by asno as the best assignment possibility color 2 The assignment given by the user is found by asno but not as the best assignment possibility color 3 The assignment given by the user is not found by asno color 4 No assignment was given by the user but asno found one or several assignment possibilities assign Es Commands c
195. r r lt T Character function that returns the character with number n Complex function that returns x ix both arguments must have the same type Complex conjugate Cosine Hyperbolic cosine Real function that returns the CPU time in seconds since the start ofthe program Character function that returns the current date in the form dd mm yy Logical function that returns 1 if a variable with name s exists and has a value different from NULL or 0 otherwise Positive difference dim x x max x x 0 Logical function that returns 1 if a variable with name s exists or 0 oth erwise Logical function that returns 1 if a file with name s exists or 0 otherwise Exponential function Character function that returns the value of the external i e non local variable with name s even if it is hidden by a local variable with the same name or a blank string if no external variable with this name ex ists Character function that returns the value of the external i e non local variable with name s even if it is hidden by a local variable with the same name or s if no external variable with the name s exists Real function that returns the Xx value of the last linear least squares fit see plot subcommand fit fiterr n fitpar n fitprob getenv s getpid global s global s s gt ichar s if N X4 Xo index sj S gt indexr s S gt int z len s A INC
196. r calculated automati cally Given a proton list called my_prot prot and a peak list called my_peaks peaks the peaks can be calibrated automatically read prot my_prot read peaks my_peaks assigned integrated caliba The simple automatic calibration is useful if no preliminary structures are available It sets the parameter A such that the average upper distance limit for the backbone calibration class becomes 3 6 A The parameters B and C are then calculated such that the calibration curves intersect at the minimally allowed upper distance limit usually 2 4 A Intersection points at higher distances would not make sense as the unphysical cal ibration functions of the type 1 7 should account for flexibility and therefore always result in a higher distance limit for the same peak vol ume 19 Tutorial 20 The calibration curves given by the automatic calibration can be refined manually For instance to tighten the calibration for the backbone cali bration class from the automatically determined value A 2 2 108 toA 1 2 108 the command caliba bb 1 2E 8 can be used If a peak volume and the corresponding upper distance limit are given the peaks of the backbone calibration class can be calibrated accordingly with the caliba macro volume 0 6E 6 d 2 4 caliba bb volume d 6 For the calibration of multiple peak lists there are two different ap proaches The first one treats every peak list sepa
197. r zero if s does not occur as a substring in s4 Integer function that returns the integer part of the real or complex num ber z Integer function that returns the number of characters in s 49 INCLAN DM length s lenstr s log z log10 z macro s match s So max x Xoy min x Xos mod x X2 mtime s nint r opened s plotx0 ploty0 plotx1 ploty1 rand 50 Integer function that returns the number of elements in the array stored in a variable with name s Integer function that returns the index of the last non blank character in sS Natural logarithm z 0 if zis real it must be positive for complex z the result has n lt Im log z lt 7 Logarithm to base 10 z 0 if z is real it must be positive for complex z the result is in the range n lt Im log10 z lt 7 Logical function that returns 1 if a macro with name s is available or 0 otherwise Wildcard match logical function that returns 1 if the string s matches the string s or O otherwise The string s may contain wildcards an asterisk matches zero or more characters and a question mark matches exactly one character Maximum Minimum Remainder of x modulo x mod x x7 x4 X int x x both arguments must have the same type x 0 Integer function that returns the time of last modification in seconds since a reference date of the file with name s Integer function that returns the
198. raints As an alternative to its native format DYANA can also read distance and angle constraint files in XPLOR format Br nger 1992 Both types of constraints are specified with assign statements followed by two or four XPLOR atom selections respectively given in free format Other statements in the input file are ignored DYANA uses a simplified version of XPLOR atom selections that supports only the resid and name ex pressions Other selection expressions and logical operators are skipped The XPLOR wildcard is converted to and the XPLOR wildcards o and are converted to In contrast to other input files com ments are started with an exclamation mark An example of an XPLOR distance constraint file is only the part printed in bold is interpreted by DYANA 165 File Formats 166 set echo false end set wrnlev 0 end assign resid 1 and name HA resid 1 and name HG 0 00 0 00 3 96 assign resid 1 and name HB resid 1 and name HE 0 00 0 00 5 50 assign resid 3 and name HN resid 3 and name HA 0 00 0 00 2 80 assign resid 3 and name HN resid 3 and name HB 0 00 0 00 3 60 assign resid 4 and name HN resid 4 and name HB2 0 00 0 00 3 60 assign resid 4 and name HN resid 4 and name HB1 0 00 0 00 2 80 assign resid 4 and name HA resid 4 and name HB2 0 00 0 00 2 60 assign resid 4 and name HA resid 4 and name HB1 0 00 0 00 2 70 assign resid 5 and
199. rans peptide bonds will be assumed through out No special covalent bond is declared The first line of the file is a comment line A special covalent bond is declared by the lowercase keyword link followed by the first atom name the first residue number the second atom name and the second residue number in free format This infor mation will only be used in DYANA to exclude the necessary atom pairs from the steric overlap check to correctly form the special covalent bond explicit upper and lower limit distance constraints are required Situa tions where special covalent bonds are needed are for instance proteins with disulfide bridges cyclic peptides or flexible proline rings To de clare for example a disulfide bridge between CYSS 3 and CYSS 55 of a protein the following entry is used in the sequence input file Ms File Formats link SG 3 SG 55 In addition the presence of this disulfide bond is then fixed directly with distance constraints e g by imposing a range of 2 0 to 2 1 A on the S S distance and of 3 0 to 3 1 on the S C distances across the bridge using explicit upper and lower distance limits Williamson er al 1985 Be cause disulfide bonds occur frequently in proteins it is not necessary but possible of course to declare them explicitly in the sequence file if the sulphur atoms of CYSS residues are not explicitly linked to other atoms by link entries in the sequence file the program allows for special co
200. rately read prot first read peaks first assigned integrated caliba read prot second read peaks second assigned integrated caliba In the second approach peak lists are read with different user defined relative weights for the peak volumes and then calibrated simultaneous ly read prot first read peaks first assigned integrated read prot second read peaks second weight 0 3 assigned integrated append caliba Because of the option append in the second read peaks command the second peak list is appended to the first peak list This approach has the disadvantage that the weights must be specified by the user Making plots 180 90 vo 90 180 180 90 0 0 90 A Tutorial Experienced DYANA users may want to create their own calibration classes or use different calibration functions Two examples illustrate this To use the uniform average model Braun et al 1981 for all NOEs involving methyl groups one first selects the corresponding peaks and then applies the uniform average calibration function weighed with the parameter C given by the user peaks select METHYL calibrate C 1 9 5 d 5 d 1 9 To calibrate all HN HN peaks with a function Ayld with the exception of the NOEs observed in or to a long and flexible loop from residue 12 to 26 which are calibrated with A d use peaks select HN 12 26 HN calibrate Al d 4 peaks select HN HN xor
201. rbars can be shown for the coupling constant values file file longrangeplot ps Plots long range distance restraints five or more residues apart versus two copies of the sequence Lines going from upper left to lower right represent restraints between side chain atoms those going from lower left to upper right represent restraints that involve backbone atoms steps N 100 dt Ar 0 05 level L taken from variable level temperature 7 0 1 accuracy e 0 0 tau t 0 0 nprint n 0 angdev Ad 10 0 vdwupdate N 4w 100 tinit z 0 0 estart T 0 01 exact continue Performs N steps of molecular dynamics in torsion angle space with step size Af including constraints up to minimization level With t 0 a molecular dynamics calculation at constant energy is per formed Otherwise the system is weakly coupled to a heat bath of tem perature T using time constant t Berendsen er al 1984 The temperature 7 can be a function 7 s of the parameter s that varies lin early from 0 to 1 during the TAD run i e in step n out of a total of N steps it has the value s n n DAN 1 If the reference value for the accuracy of energy conservation e has a positive value the length of the integration time step Ar will be adapted 99 Commands 100 during the run in the same way as the temperature such that the relative change of the total energy in successive integration steps is close to In this case the para
202. re deleted When working with angle constraints all constraints for angles whose name matches one of the angle specifications on the command line are deleted Switch to DIFIMA the part of the program for the manipulation of dis tance constraint files macro Give a directory of all standard macro files and all macro files in the cur rent working directory If a macro specification is given the directory will only contain those macro files with names that match the given mac ro specification A macro specification is a macro file name possibly containing wildcard characters but excluding the extension cfm For every macro its name and the comment lines that precede the first com mand line are listed Aj A range Remove the covalent connectivities of the specified atoms This com mand can only be used with coordinate files The default is to remove all covalent connectivities distances extract help A COFIMA A range A range condition When working with Cartesian coordinates calculates the distances be tween atoms specified by A range and atoms specified by A rangen When working with distance constraints list constraints for distances between atoms specified by A range and atoms specified by A rangen Optionally only distances or constraints that fulfill one or several of the following conditions are listed d lt value distance less than value d
203. reak command do do 54 prompt variable Writes the string prompt to standard output reads one line from standard input and assigns from this line strings separated by blanks to the given variables The command is usually used for interactive input within mac ros A prompt that contains blanks must be enclosed in double quotes ask First and last point begin end First and last point 12 45 print range begin end range 12 45 Breaks a do loop and is only allowed in macros The execution of the macro is continued with the first statement following the loop name Defines a new globally visible user defined command within a macro 1 e a macro within a macro User defined commands defined by com mand statements are called by their name possibly followed by param eters in exactly the same way as macros Within a macro a user defined command can only be called after it was defined The statement com mand without parameters gives a list of all user defined commands and indicates where they are defined without parameters Executes a loop within a macro The loop is exe cuted unconditionally i e until one of the statements break exit quit or return is encountered do if filename eq break end do variable start end step parallel continue Executes a FORTRAN 77 do loop within a macro The loop counter vari able and the integer expressions sta
204. reates the standard upper and lower limit distance constraints Will iamson et al 1985 to enforce a hydrogen bond between two atoms one of which must be a hydrogen atom The distance between the hydrogen and the acceptor is restrained to the range 1 8 2 0 A and the distance be tween the atom covalently bound to the hydrogen and the acceptor is re strained to the range 2 7 3 0 Contains commands that are executed automatically at the start up time of DYANA e g the setting of important variables and the definition of some aliases After the general init macro a user defined init macro in the current directory is executed 1f available dist 0 0 Keeps only assignments with a reliability distance gt dist The reliability distances must have been calculated previously with the command reli ability ile file kringle ps delta A 30 errorbars longrangeplot md Es Commands Produces a graphics output file with the given name a GRAF file if the extension is grf a MIF file if the extension is mif or a Postscript file otherwise containing a plot of Jaga versus Tapa coupling constants Nagayama Wiithrich 1981 The theoretical curve based on the Kar plus equation given in the library is also drawn both for a rigid structure solid line and for the situation when the x angle is uniformly distrib uted in the interval ZA around a given value dotted line Optionally er ro
205. red norm of the gradient of the target function is smaller than the value of the parameter GSQTOL in the subroutine CGMIN maxit The maximal number of target function evaluations has been exceeded linmin The maximal number of target function evaluations dur ing the line minimization see the parameter MAXLIN in 101 Commands noah 102 the subroutine CGMIN has been exceeded without de creasing the target function value nostep The step size during line minimization became too small uphill The direction of a conjugate gradient minimization step was uphill const Several conjugate gradient steps did not succeed in de creasing the target function see the parameter MAX CON in the subroutine CGMIN flat The target function was minimized by less than 1 dur ing the last n iterations stuck Several attempts to restart the conjugate minimizer after an update of the list of steric constraints failed If 64 bit floating point precision is used normal stop criteria are gradtl maxit and flat All others should occur only rarely With 32 bit floating point precision other stop criteria may occur due to non serious numer ical problems num_cyc n 24 peak_namsjile1 file2 plformat string string2 determined by peak list file rmsd_range residue range all residues protein name proton_nam 7jile file2 minimizer macro noahmin peak_ref file1 file2 options string addupl file
206. rod lge Tet lle and Ut are available by their generic names but not un der special names For example the absolute value function is known by the name abs but not by the special names iabs or cabs e There are additional intrinsic functions see below e Blanks can only appear at reasonable places but not inside of num bers variable names etc Intrinsic functions In the following list of all INCLAN intrinsic functions arguments are de noted by n integer r real c complex sS string x integer or real unless types are given explicitly z real or complex The result type of an intrinsic function is only given explicitly if it dif fers from the type of the argument s Absolute value the argument x is of any numeric type for complex ar guments the result is real Arc cosine r lt 1 0 lt acos r lt m Real function that returns the imaginary part of c Discard fractional part the result if of type real Closest integer the result if of type real Arc sine r lt 1 7 2 lt asin r lt 1 2 Arc tangent n 2 lt atan r lt 71 2 47 INCLAN DM atan2 r o char n cmplx x1 X2 conjg c cos Z cosh r cputime date def s dim x x2 exist s existfile s exp 2 external s external s S gt fitchisq 48 Argument of the complex number r ir not r ir r and r must not both be zero T lt atan2
207. rposes are written Optionally this variable may have two of the above values separated by a comma In this case the first value applies to standard output the sec ond to the protocol file denotes the number of command line parameters of the current macro denotes the maximal number of processors that is used for parallel do loops are the default names for the command line parameters of a macro These names may be changed at the beginning of the macro 45 INCLAN path prompt protocol timing 46 denotes the search path for macro files in the form of a comma separated list of directories denotes the prompt for interactive input If this variable is not defined or blank no prompt is written but multiple blank lines of input and the end of the execution of a macro are indicated by the word Ready on a sep arate line denotes the name of the protocol file into which standard output is dupli cated under the control of the variable info If this variable is not defined or blank no protocol file is written is a system variable to control the reporting of CPU times CPU times are given for all commands except for those that are built into the command line interpreter that need more seconds of CPU time than the value of timing indicates Expressions The command interpreter can evaluate general FORTRAN 77 integer re al complex logical and character expressions Expressions can appear in eval stateme
208. rt end and step have the usual mean ing Parallel loops are executed in parallel on nproc processors If the keyword continue is present the program continues immediately with the execution of the next statement after the parallel loop Otherwise the else else if end end do end if error eval A INCLAN next statement after the loop is executed when the parallel loop is fin ished do i 1 10 print Iteration i end do Starts an else clause of a block if statement condition then Starts an else if clause of a block if statement Ends a user defined command or subroutine Ends a do loop Ends a block if statement text Writes the text to standard output or into the file with the given filename and calls the error handler This statement is suitable to treat errors that occur during the execution of a macro If the text contains blanks it must be enclosed in double quotes variable expression Evaluates the arithmetic or string expression according to the rules of FORTRAN 77 and assigns the result to the variable The keyword eval can be omitted In contrast to FORTRAN 77 function names must be given in lowercase letters eval i 7 sentence A flexible program j mod i 4 2 1 len sentence show i sentence j 1 Variables i 7 sentence A flexible program 55 INCLAN external exit go to 56 324 J 9 1 19 variable expr
209. s e noah ps PostScript file with summary of the NOAH run Ds Tutorial e noah grf Graf file for noah ps containing RMSD and target func tion values after each cycle as well as the number of assigned peaks e noah x peaks Assigned peak lists lt incomp x peaks Peak list with peaks that are incompatible with the final structure bundle e reliability x Reliability distance of every assigned peak e diff x Differences in the assignment compared to the reference peak list only if such a peak list was indicated with the parameter peak_ref e end upl end lol Upper and lower limit distance constraint files used for the final calculation 66 99 x is the peak list number x 1 n The file noah ps is gener ated by INCLAN using the noah grf file which contains all numbers RMSD target function values and assigned peaks Range for RMSD calculation 3 37 Cycle RMSD all RMSD bb TF 1 TF 10 Da 64 32 5 25 161 3 190 3 6418 9 11 120 0 159 8 2 6 30 5 36 126 9 1545 3 6 06 5 13 12 745 160 0 4 5 40 4 29 8 9 24 5 37 9393 A 82 104 4 136 7 6 5 65 4 71 100 1 146 8 Ti 2949 4 52 108 9 146 3 8 4 55 3 52 12 7 21 59 20 94 1 34 30 6 1 21 7 La 0082 Sd 6 5 22 37 0 89 4 6 7 0 23 39 0 93 2 1 4 8 25 46 0 92 1 6 2 9 a Number of assigned peaks Total new different 1 372 21 0 416 26 0 433 29 0 441 32 1 449 33 1 501 39 1 505 40 1
210. s helix lol and dihedral angle constraints helix aco for a 11 res idue polypeptide the second helix of a mutant of the Antennapedia ho meodomain from Drosophila melanogaster G ntert et al 199 1b Output from the program is indented for example the confirmation that the sequence was successfully read Sequence file helix seq read 11 residues The following command calculates a group of 5 conformers dyana gt calc_all 5 5 random structures created seed 3771 Structure annealed in 2 s 0 630422 Structure annealed in 2 s 0 112963 Structure annealed in 2 s 0 197074 Structure annealed in 2 s 0 314798 Structure annealed in 2 s 0 384202 5 structures finished in 11 s 2 s structure First 5 structures with random values for the dihedral angles are created Then for each of these random structures violations of the conforma tional constraints are minimized by simulated annealing using torsion angle dynamics G ntert ef al 1997 Cartesian coordinates of all resulting structures can be saved with the write cor command dyana gt write cor helix all DG coordinate file helix cor written 5 conformers Acomplete protein structure calculation Ds Tutorial To get an overview of the quality of the minimized structures the over view command can be used dyana gt overview helix This example provides a complete prototypical prote
211. s between the two residues or residues close to them that support the given distance constraint A score of zero indi cates that there is no other NOE observed between the two regions of in terest This is not only very suspect but also quite dangerous because such a single long range NOE may have dramatic effects on the struc ture All peaks which cause such dangerous long range distance con straints should be checked manually directly in the spectra In the following example the long range NOE from residue 3 to 62 has a score of 0 0 because there is absolutely no other NOE supporting it while it seems unlikely that all the NOEs observed between residue 6 and 57 are derived from wrong assignments dyana gt distance check Distance constraint Score Upper QE LYS 3 s0D LYS 62 0 00 Upper HA TYR 6 OB ALA 53 2 00 Upper HB2 TYR 6 OD1 EU 57 SW ES Upper HB3 TYR 6 OD1 EU 57 3 18 Upper OB TYR 6 OB ALA 53 2 00 Upper OB TYR 6 OD2 EU 57 4 25 Upper QD TYR 6 HA ALA 53 22 90 Upper OD TYR 6 OB ALA 53 1 50 37 Tutorial 38 INCLAN The user interface of the program is based on INCLAN a powerful inter active command language that allows the use of variables FORTRAN 77 mathematical and character expressions macros flow control loops conditional statements jumps the production of graphics etc When reading an input command line the command interpreter executes the follo
212. sidues angles side chain angles CHI1 t cut fmax 0 0 continue Performs for all amino acid residues in the given range grid searches comprising the backbone dihedral angles 6 w and the given side chain angles To specify more than one side chain angle the names must be given separated by blanks and enclosed in double quotes If the cutoff value for the local fragment based target function fmax is positive then all conformations with a local target function value below fmax Will be considered as allowed Otherwise a conformation will be allowed if no single restraint violation exceeds the corresponding cutoff value defined by the variables soft_upl soft_lol etc Unless the continue option is set the allowed ranges of dihedral angles will be initialized to allow all possible angle values before the grid searches are started 97 Commands hbond init keep kringle 98 This macro provides the functions of the former HABAS program G n tert et al 1989 The results include dihedral angle restraints and if pos sible stereospecific assignments for the diastereotopic groups in the fragment habas angles CHIl CHI2 tfcut 0 05 Perform grid searches for all amino acid resi dues including the dihedral angles 6 y x and x Allow conformations with local target func tion values up to 0 05 atom1 atom name a residuel residue number atom2 atom name residue2 residue number C
213. specification is omitted the previously used output file is used Sequences of commands that are often used may be stored in macros different from INCLAN macros with file name extension cfm in order to facilitate routine applications of the program Macros can be called from within a macro When executing a command the program can detect two different types of problems warnings which cause only the current command to be skipped and errors which cause the whole rest of the macro to be skipped Macros can be commented text between the comment sign and the end of a line is considered as a comment A set of standard macros is provided with the program am_di Change from AMBER to DYANA nomenclature am_fm Change from AMBER to FANTOM nomenclature angles A COFIMA am_op Change from AMBER to OPAL nomenclature attach_am Attach hydrogens to amino acids and DNA AMBER con ventions backbone Keep only backbone atoms N CA C di_am Change from DYANA to AMBER nomenclature di fm Change from DYANA to FANTOM nomenclature di op Change from DYANA to OPAL nomenclature di Kp Change from DYANA to XPLOR nomenclature fm_am Change from FANTOM to AMBER nomenclature fm_di Change from FANTOM to DYANA nomenclature fm_pdb Rename residue names and last atom from FANTOM to PDB fm_xp Change from FANTOM to XPLOR nomenclature heavy Keep only heavy atoms norm_residues Achieve standard three letter code for amino acid res idues
214. st and second atom respectively the distance limit in and optionally the relative weight of the constraint The default relative weight is 1 Relative weights should be positive The weight of a con straint in the target function equals the relative weight times the weight 163 File Formats I ing factor for the corresponding type of constraints An example file follows 29 ARG HN 29 ARG HB2 2 90 29 ARG HN 29 ARG HB3 3 00 29 ARG HN 29 ARG QG 4 33 29 ARG HN 30 ARG HN 3 30 29 ARG HA 29 ARG QG 3 87 29 ARG HA 32 ARG HN 4 00 29 ARG QD 33 ILE OD 6 80 30 ARG HN 30 ARG OB 2 99 30 ARG HN 31 ARG HN 3 40 30 ARG HN 33 ILE CB 9 10 30 ARG HA 30 ARG OB Pb 30 ARG HA 30 ARG OD 5 80 30 ARG HA 33 ILE HN 3 80 5 00E 00 In this example the last constraint has a relative weight of 5 all others have the default relative weight of 1 If on an input line the first residue number and name are absent the corresponding data from the previous data line is used On the other hand the first residue number and name may stand alone on a line such that the following is an equivalent form of the above example distance constraint file 29 ARG HN 29 ARG HB2 2 90 HN 29 ARG HB3 3 00 HN 29 ARG QG 4 33 HN 30 ARG HN 3 30 HA 29 ARG QG 3 87 HA 32 ARG HN 4 00 QD 33 ILE QDI 6 80 30 ARG HN 30 ARG OB 2 99 HN 31 ARG HN 3 40 HN
215. st be an integer real complex logical or character con stant according to the rules of FORTRAN 77 In particular character strings must be delimited with single quotes Variables can be used in both ways simultaneously which makes them a powerful tool of the command language A variable name consists of up to 32 letters digits or underscore char acters _ The value of a variable is always stored as a character string and only converted temporarily to an integer real or complex number during the evaluation of a FORTRAN 77 expression There are several types of variables 41 INCLAN DM Local variables Global variables Special variables System variables Basic substitutions Fortran format 42 exist only within the macro where they are declared and in macros called from this macro With the exception of the command line param eters of a macro which are always local local variables must be declared in var or syntax statements They exist until they are removed with un set or until the end of the macro in which they are declared is reached are always visible except when they are hidden by local variables with the same name Variables that are not local are global The user can in troduce new global variables simply by using a variable with a new name Global variables exist until they are removed with unset are variables that can be created and used by the user but have also a spe cial meaning to the
216. structures Lists all selected peaks factor f Scales the volumes of the selected peaks by the factor f peak selection Selects all peaks that match the given peak selection see chapter Selec tions dim d all dimensions Deletes the assignment of the selected peaks Optionally only assign ments in dimension d may be deleted average maximum average 105 Commands A ramachandran random _all randomize read aco 106 From each group of identically assigned peaks only one peak is kept Peaks are considered as identically assigned if they are assigned to the same proton pair For instance in a2D NOESY spectrum a cross peak and its transposed peak are identically assigned Peak volumes may be averaged or their maximum be kept Only peaks with positive volume are considered in the averaging procedure file name ramachandran ps nobackground label Produces a graphics output file with the given name a GRAF file if the extension is grf a MIF file if the extension is mif or a Postscript file otherwise with a Ramachandran plot of the selected structures The background consists of three different blue tones indicating most fa vored regions dark blue additional allowed regions medium blue and generously allowed regions light blue It corresponds to the background found in the program PROCHEC
217. t and angle define the font used to write the text In addition the text may contain the following embedded text com mands T Change font type to Times H Change font type to Helvetica C Change font type to Courier S Change font type to Symbol b Change to boldface i Change to italics A Start a superscript v Start a subscript N Return to standard font end sub or superscript If the text contains multiple blanks it must be enclosed in double quotes Double quotes that are part of the text must be preceded by a backslash See section mark text writes text into the output plot file Plot parameters are used to define the positioning and appearance of graphics objects They are set by the plot subcommand set determines how text is aligned with respect to its reference position Pos sible values are left The horizontal reference position is at the left margin of the text center The horizontal reference position is in the center of the text right The horizontal reference position is at the right margin of the text bottom The vertical reference position is at the bottom margin of angle autoscale border color A INCLAN the text middle The vertical reference position is in the middle of the text top The vertical reference position is at the top margin of the text Horizontal and vertical alignment specifications can be separated by a comma e g align center top
218. te upl caliba upl save upper limits before modifications Calibration is performed separately for each peak list using the default method implemented in the macro caliba See the section Calibrating NOEs later in this tutorial for more details on calibration 11 Tutorial 12 The next step is a systematic analysis of the local conformation around the C atom of each residue using grid searches On the basis of local distance constraints from the file caliba upl and scalar coupling con stants from the file er2 cco allowed conformations for the dihedral an gles 6 y x and x of each residue are determined using the macro GRIDSEARCH read seq name seq read sequence and initialize read upl caliba upl read NOE upper distance limits read cco Sname cco read scalar coupling constants atoms stereo HB2 2 5 atoms stereo QD1 39 Define stereospecific assignments that are al ready known habas angles CHIl CHI2 tfcut 0 05 Perform grid searches for all amino acid resi dues including the dihedral angles 6 y x and x Allow conformations with local target func tion values up to 0 05 A2 gridplot habas ps create plot s with allowed angle ranges atom stereo list List stereospecific assignments distance modify modify distance constraints write upl name upl save upper limits write aco name aco save angle restraints The command distance modify removes irrelevant constraints con straints that involve fixed distan
219. ted 20 structures 115 Commands sugarbond sugarring translate vtfmin 116 range residue range all nucleotides Defines the correlations between the dihedral angles and the pseudoro tation angle Pin DNA RNA sugar rings residue number residue of current fragment Creates 5 upper and 5 lower limit constraints to close the bonds be tween C4 and O4 in the ribose rings of the nucleotides in the given res idue range xplor on off clear Defines atom and angle name translations between the nomenclature used in the standard DYANA residue library and other commonly used nomenclature systems This allows reading of input files and writing of output files according to other nomenclature systems Currently xplor nomenclature is supported The options on off and clear have the same meaning as in the atom rename and angle rename commands Without option translate lists the currently set atom and angle name translations translate xplor Set XPLOR translation table read xplor noe tbl Read a XPLOR distance constraint file read pdb in pdb unknown warning Read a PDB file with XPLOR nomenclature Avoid errors if unknown atoms are encountered translate off Use standard DYANA nomenclature again translate on Switch to XPLOR nomenclature write pdb out pdb Write a PDB file with XPLOR nomenclature levels L L gt O number of residues steps N N gt 15
220. that are covalently bound to atom n Number of coupling constant constraints Number of allowed conformations in the most recent grid search Total number of dihedral angles free and fixed Number of distance constraints between residues that are exactly n posi tions apart in the primary sequence Number of distance constraints Number of distance constraints that involve the residue n and span a dis tance of at least m positions in the sequence Number of free i e rotatable dihedral angles Number of distance constraints between residues that are less than n po sitions apart in the primary sequence Number of lower limit distance constraints NOAH variable Number of peaks in the unambiguous assignment list af ter the command filter NOAH variable Number of peaks in the unambiguous assignment list that have the same assignment as in the reference peak list after the com mand filter NOAH variable Number of peaks that are incompatible with the current structures after the command assign NOAH variable Number of peaks in the unambiguous assignment list that have no assignment in the reference peak list after the command fil ter np_out np_wrg npeaks nplist nr nseldis nstruct numpro n nupl pi pseudoatom n rad rmsd_bb rmsd_bbdev rmsd_hv rmsd_hvdev rnam n rnum n A Variables and Functions NOAH variable Number of peaks that have no possible assignment based on
221. that excludes the torsion angle values of these regular secondary structure elements Torsion angle re straints for x are depicted by filled squares of three different de creasing sizes depending on whether they allow for one two or all three of the staggered rotamer positions 60 60 180 Tor sion angle restraints for X that exclude all three staggered rotam er positions are shown as filled circles Upper distance limits for sequential and medium range distances are shown by horizontal lines connecting the positions of the two residues involved The thickness of the lines for the sequential distances dyn i i 1 dyn i i 1 and dgn i i 1 is inversely proportional to the squared upper distance bound This command should be executed before distance modify because many of the intra residual and short range distance constraints will be removed by distance modify because the do not effectively restrict the conformation Rao Creates the standard upper and three lower limit distance constraints Williamson er al 1985 to enforce disulfide bonds between pairs R Ro R3 Ry etc of cystine residues These residues must be of the type 113 Commands gt stereoassign structure clear structure copy 114 CYSS For a disulfide bridge between residues i and j three upper lim its and three lower limits are generated 2 0 lt d S SY lt 2 1 3 0 lt a Ch SY lt 3 1 6 3 0 lt d S C
222. the data lines is 5X 3A5 3F8 2 Residue sequence The sequence input file defines the primary structure of the molecule un der consideration identifies residues following cis peptide bonds de clares special covalent bonds i e covalent bonds that are not compatible with the tree structure of the molecule and identifies fixed and rotatable dihedral angles in the molecule The residue names consist of up to four characters and must of course match the name of a residue entry in the residue library file If a residue name is preceded immediately by a lowercase c the dihedral angle of the peptide bond preceding this residue will be fixed at cis position in stead of trans position throughout the calculation Optionally a residue name may be followed by its residue number by default the residue number of the first N terminal residue in the sequence is set to one and for other residues the residue number will be the residue number of the preceding residue plus one Different residue names and numbers must be separated by at least one blank or end of line character otherwise the format is free The syntax to declare fixed and rotatable dihedral angles is explained below An example sequence input file follows Second helix of Antennapedia Homeodomain ARG 29 ARG ARG ARG ILE GLU ILE ALA HIS ALA LEU This file contains the sequence of a peptide that is 11 residues long with residue numbers 29 39 T
223. the first row shows a positional pa rameter the second row shows a named parameter the third row shows options that may be given simultaneously and the last row shows mu tually exclusive Options see INCLAN command syntax Sometimes optional items are given in square brackets and indicates that the preceding item may be repeated several times When executing a command the parameters and options must all be giv 75 Commands A angle fix angle flip angle free angle list angle rename 76 en on one line or on continuation lines the tabular form is used only in the manual for clarity angle selection all angles All selected angles are fixed i e become non rotatable and cannot be changed during minimization or dynamics By default all peptide angles are fixed and set to 180 see also Residue sequence in chapter File formats angle selection all angles All selected angles of the selected memory structures are analyzed to find the most frequent angle value The angles are then flipped by 180 1f the new angle is nearer to this most frequently found angle value This command is used by the flip macro angle selection all angles All selected angles become free angles 1 e rotatable angles that may be changed during minimization or dynamics see also Residue sequence in chapter File formats angle selection all angles
224. the points x y The total number of points must be 3n with integer n22 lx yy list mode draws shapes enclosed by B zier spline curves through the points of the given vector expressions x yj If no vector expressions are specified shapes are drawn for all list columns If the x expression is omitted i e if only a single expression y4 is given the x coordinates are taken from the x column of the list The number of list points must be 3n with inte gern KIYI 2 2 normal mode draws a cubic spline through the points a Vj X Yo The spline starts at the first point and ends at the last point with vanishing second derivative The x values must be increasing x lt x for all i lx y list mode draws cubic spline curves through the points of the given vector expres sions X yj If no vector expressions are specified splines are drawn through the points of all list columns If the x expression is omitted i e if only a single expression Vj is given the x coordinates are taken from the x column of the list See section mark 69 INCLAN text triangle write align 70 x y text print text at position x y The alignment of the text with respect to the reference position x y depends on the current values of the plot pa rameters align and rotate The current values of the plot parameters font textsize weigh
225. the vector expressions x y and O respectively For m basis functions f fm the optimal linear combination y x af tan km 1 is determined by minimizing AIN ia re 2 where i runs over the list data points The optimal fit function y x is added as another column to the list data This command does not draw anything The fit parameters a 4 their standard deviations x and the probability that this value of x would be exceeded by chance are available through the intrinsic functions fitpar fiterr fitchisq and fitprob respectively If the errors of the data points are unkown this can be indicated by setting to zero in the fit command dot x yl Plot original data points fit x log y1 01x Logarithmic fit of y a exp a x spline x exp y2 Plot fitted curve 65 INCLAN frame function label line 66 xtext xtext ytext yrext ticS ticaxes X Y labels abelaxes X Y grid zero draws a rectangular frame with corners X Yo Ng Y X Yo and X Y Subsequently produced graphics is clipped on the bor ders of the frame The x and y axes are labeled with the titles xtext and ytext respectively The parameter tics and labels determines whether tics and numeric labels are drawn The possible values for ticaxes and la belaxes are off No labels or tics x Labels or tics only on the x axis y Labels or tics only on the y axis X Y Label or tics
226. three parts COFIMA for coordinate file manip ulations DIFIMA for distance constraint file manipulations and ANCOMA for angle constraints file manipulations The prompts cofima gt di fima gt and ancoma gt indicate the part of the program that is current ly active Many commands can be used for all three types of data files but there are also commands that are specific for certain types of data files Commands can be abbreviated as long as the abbreviation remains unambiguous In the following description of the individual commands A Ay etc de note atom or angle names R R etc denote residue names and r r etc denote residue numbers Atom angle and residue names must start with a letter and may except in some cases contain wildcards stands for zero or more arbitrary characters stands for exactly one arbitrary character No blanks are allowed within names Residue numbers are in tegers Atom angle residue names and residue numbers may be preced 171 COFIMA 172 cepo ed by an exclamation mark which acts as a not operator The special residue names FIRST and LAST can be used to denote the first and the last residue in the coordinate file respectively The special resi due names first and last can be used to denote the first and last residue of every fragment with contiguous residue numbers in the coordinate file respectively Atom angle and residue names but not co
227. tions three sub cat egories can be made Mumenthaler et al 1997 e RelDis 0 0 A MinVio 0 0 A Both assignments are satisfied in the structures Assuming that the conformers are correct solutions such peaks must be superpositions of two NOE signals so that both 87 Commands A dinucleotide distance check 88 assignments are correct e RelDis gt 0 0 MinVio 0 0 A Here the assignment from the peak list file lies outside of the given tolerance range from the peak position and was therefore not considered during the calculation of the reliability distance Unless the current proton shifts are not well adapted to the peak list or the tolerance range was too small the cur rent assignment seems more appropriate for the peak under consid eration e RelDis gt 0 0 MinVio gt 0 0 A These are the most relevant differ ences since the assignment from the peak list file is violated by the current structures Thus the different assignment will also have an impact on the structures If the assignment in the peak file has a sign in the integration method field the comment Peak not used in structure calculation is written Optionally an intersection is made between both peak lists and the as signments are kept only if the peak is assigned to the same proton pair in both lists If the peak is assigned only in one of the lists it is therefore also unassigned The option notdiff is less stringent sinc
228. to start the programs will be created and in stalled in the directory bin directory by cd install directory dyana 1 5 setup bin directory The default bin directory is HOME bin where HOME is the value of the corresponding UNIX environment variable Ms Installation To have easy access to the program DYANA the bin directory should be included in the search path of the UNIX shell If DYANA was correctly in stalled and the directory containing the executables is included in the UNIX search path then the programs can be started simply by typing their name If the program DYANA can be started but does not display the prompt dyana gt the installation is not correct and should be repeated To remove all files created by make the command make clean may be used After make clean it will be necessary to run the configu ration script again After copying the program from one computer to an other it is important to perform make clean before running the configuration script Similarly make uninstall removes all files created by make install and make recompile completely recompiles the programs but preserves executables for other computer types that are already present 185 Installation Da 186 References Bartels C Xia T Giintert P Billeter M amp Wiithrich K 1995 The pro gram XEASY for computer supported NMR spectral analysis J Biomol NMR 5 1 10 Berendsen J H C Postma J P M
229. tored in a GRAF file i e a file with extension grf called curve grf and supplemented with the ap propriate GRAF commands 1 8 27 10 0 54 frame line This GRAF file can be converted into in a Postscript file curve ps with the command graf curve Similarly a histogram of the same data can be produced by replacing the last line of the file curve grf with fi11 1 rectangle x 0 2 0 x 0 2 yl fill 1 selects pattern 1 solid to fill the interior of polygons and rect angle x 0 2 0 x 0 2 yl has the meaning plot for every data point x y in the table of values a rectangle with lower left corner x 0 2 0 and upper right corner x 0 2 y 23 Tutorial DI A4 Using INCLAN variables 24 In the above examples the plots had the default size and position and the scales for the x and y axes have been chosen automatically by GRAF However the user can also specify these parameters explicitly The positioning of a plot on the paper is governed by the four parameters x0 yO x1 and y1 that specify the positions of two reference points Xp Yo and x y1 in a coordinate system centered in the middle of an A4 sheet with axes running in the mathematically usual directions and using typographical points 1 pt 0 353 mm as units By default the two reference points specify a large square placed in the center of an A4 sheet xy Yo 250 and x y 25
230. ue entries A residue entry starts with a header line with the Fortran format A10 A5 415 and containing the word RESIDUE the residue name the numbers of rotatable dihedral angle and atom declarations that will fol low respectively and the numbers of the first and last atom in the list of atom declarations that belong to the residue not to the preceding or fol lowing one in the polypeptide chain The next lines contain dihedral an gle declarations in the format 5X A5 20X 515 the dihedral angle name the numbers of the four atoms that define the dihedral angle and Atom types ALA Ds File Formats the number of the last atom that will be affected by a rotation of the di hedral angle for backbone dihedral angles this number is set to 0 Atom numbers correspond to the running numbers in the first column of the atom declarations The atom declarations must be ordered such that the set of atoms affected by a change of a dihedral angle consists of all atoms following the third atom in the dihedral angle definition up to the last atom or the C terminus for backbone dihedral angles that is affected Finally there are lines containing atom declarations the format is 5X 2A5 15X 3F10 4 515 the data are the atom name the atom type used to set the repulsive core radii the x y and z coordinates in for an arbitrary conformation four atom numbers indicating covalent con nectivities if there are less than four connectivities th
231. umber of dimensions 2 4 3 339 10 048 1U 1 183e 05 0 00e 00 e 0 28 23 overlap 5 2 791 10 048 1 U 3 090e 05 0 00e 00 e 0 27 23 transposed 6 6 307 9 858 1 U 1 810e 05 0 00e 00 e 0 46 44 Wi 3 179 9 858 1 U 3 506e 04 0 00e 00 e 0 49 44 8 4 570 9 939 1 U 0 000e 00 0 00e 00 0 67 65 9 4 361 9 939 1 U 2 429e 03 0 00e 00 e 0 2420 65 10 1 226 9 939 1 U 1 793e 05 7 5le 01 d 0 2421 65 The first two peaks carry comments overlap and transposed re spectively The integration method code is either e for peaks that have been integrated or for peaks that have not been integrated An example of a three dimensional peak list is Number of dimensions 3 45 52 530 4 738 4 738 1 1 903e 05 6 88e 03 a 0 22 23 23 46 52 530 3 024 4 738 1 1 842e 04 8 93e 02 a 0 22 25 23 47 52 530 2 956 4 738 1 3 620e 04 1 02e 03 a 0 22 26 23 51 38 080 8 635 3 024 1 6 100e 04 2 70e 02 a 0 24 21 25 52 38 080 4 738 3 024 1 1 872e 04 2 73e 02 a 0 24 23 25 53 38 080 3 024 3 024 1 2 776e 06 1 25e 04 a0 24 25 25 54 38 080 2 956 3 024 1 2 922e 06 1 45e 04 a 0 24 26 25 55 38 080 7 400 3 024 1 1 155e 05 2 89e 02 a0 24 28 25 56 38 080 6 660 3 024 1 2 673e 04 3 01e 02 a 0 24 29 25 Upper and lower distance limits The upper and lower distance limit files are used to enter distance con straints into the program DYANA For each distance constraint there is a line with the following data residue number residue name and atom name of the fir
232. unction maxang In general a structure is stored in the form of all dihedral angle values However some commands e g rmsd require direct access to the Cartesian coordinates of the struc tures For this reason for a limited set of structures both the dihedral an gle values and the Cartesian coordinates are stored The maximal number of structures for which Cartesian coordinates can be stored is given by the function maxcor In a DYANA calculation most operations e g minimization are per formed on the current structure The structure copy command can be used to save the current structure 17 Tutorial DI Handling stereospecific assignments 18 do i 1 10 Loop over 10 structures random Create random structure anneal Apply simulated annealing protocol to current structure structure copy 0 i Copy current structure to structure end do Structures can then be sorted by their target function values with the command structure sort and listed with structure list Structural statistics str target upper limits lower limits van der Waals torsion angles function sum max sum max sum max sum max 1 0 12 0 0 7 0 14 0 0 1 0 09 O 0 4 0 13 0 0 0 0 01 2 0 49 1 1 5 0 33 0 0 0 0 03 0 23 A0K25 0 0 1 0 06 3 0 28 T Li z 20827 0 0057 10517 0 207 70513 0 0 1 0 04 4 06427 Or Led 20618 000 03 Mil 0 12 0415 0 0 1 0 12 5 0 44 0 1 2 0 20 0 0 1 0 10 0 1 4 0 20 0 0 1 0 03 Ave 053510 2 2 0 21 0 40 2 0 dd 0
233. us angle correlate command then the new angle will be correlated by the function f p to the same parameter p as the old angle grid fragment DELTA NU1 NU2 4 Define a molecular fragment consisting of the sugar ring of nucleotide 4 in DNA or RNA numax 40 0 rad amplitude rad 180 7 grid correlate numax cos p 2 pi 5 2 pi 3 DELTA grid correlate numax cos p 2 pi 5 DELTA NU1 grid correlate numax cos p DELTA NU2 Correlate the dihedral angles in the sugar ring to the pseudorotation angle angle selection all angles none Defines a fragment to be analyzed by a subsequent grid search The an gle selection must select a connected subset of the dihedral angles Al ternatively the option none can be given to undefine the current fragment grid fragment PSI 7 PHI PSI CHI1 8 PHI 9 Defines a molecular fragment consisting of y of residue 7 6 wand y of residue 8 and 6 of res idue 9 95 Commands A grid memory grid search 96 grid memory expression none angle selection all angles in fragment This command handles the storage of allowed dihedral angle values that have been determined by grid searches These are stored in grid memo ries that contain for each dihedral angle in the molecule not only in the current fragment a fine grid of 2 spacing to store the allowed values The standard grid memory A is used by the grid search commands oth er grid memories with user define
234. ut parameters consists of 4000 TAD steps One fifth of these are performed at an initial high temperature fol lowed by slow cooling during the rest of the schedule Various parame ters of the standard annealing protocol can be changed by the user For instance 6000 TAD steps will be performed with the command anneal steps 6000 An ensemble of structures can be calculated using TAD with the macro calc_all With Calculating structures using the REDAC strategy Ds Tutorial calc_all 30 30 structures are calculated by applying the standard protocol anneal to 30 start cconformers with random torsion angles The resulting conform ers are stored in structure memories 1 30 To use instead of anneal an other modified annealing schedule that is say stored in a macro myanneal the command is calc_all 30 myanneal steps 5000 steps 5000 is an example of a parameter that will be passed to the myanneal macro An overview file helix ovw an angle file helix ang and a coordi nate files helix cor of the 20 conformers with lowest target function value can be generated after the structure calculation with the command overview helix structures 20 ang cor Molecular dynamics in torsion angle space is the preferred structure cal culation method for all proteins except maybe small helical peptides In the REDAC strategy G ntert amp Wiithrich 1991 an ensemble of n structures is first calculated with the var
235. valent bonds between all such sulphur atoms of CYSS residues i e in the van der Waals check it potentially allows disulfide bridges between any two CYSS residues in the molecule Another frequent case where the program generates a special covalent bond implicitly occurs if the bond is present in the list of covalent connectivities of the atom entries in the library but not compatible with the tree structure of dihedral an gles This is the case for example in flexible sugar rings of the DNA Nevertheless explicit upper and lower limit distance constraints are still required to enforce correct bond lengths and angles By default the program DYANA assumes that all dihedral angles de clared in the residue library are rotatable i e are degrees of freedom during the minimization The only exception are angles called OMEGA which are by default fixed at 180 or 0 To obtain different choices of fixed and rotatable dihedral angles angle declarations have to be inserted into the sequence file To make a dihedral angle rotatable use the syntax angle free where angle stands for the angle name To fix a dihedral angle at the value of the input conformation that will be read use an gle fixed This type of declaration cannot be used if the start confor mations are generated randomly within the program To fix a dihedral angle at a given value use angle value The value has to be given in degrees The angle name may contain wild
236. ven color Initial value black 71 INCLAN MI dash fill LI E HAH LI ZANIERAN font linewidth marksize mode 72 defines the dash pattern used to draw lines Its value is either blank which is equivalent to solid or a comma separated list of numbers or one of the following literals solid Solid lines dotted Dotted lines equivalent to 1 dashed Dashed lines equivalent to 5 4 dot dashed Dot dashed lines equivalent to 5 2 1 2 General dash patterns are specified by a comma separated list of num bers that define the lengths measured in points of alternating solid and invisible stretches Initial value solid defines the fill pattern used to draw areas Its value is an integer between 0 and 15 with the following meaning 0 Empty do not fill areas 1 Full color 2 7 Progressively less saturated shading or color 8 White covers other graphics 9 15 Different types of hatching Initial value 0 defines the font type and can have the following values Times Times Helvetica Helvetica Courier Courier Symbol Symbol Initial value Helvetica defines the current linewidth in points 1 pt 0 353 mm Initial value 1 defines the mark size in points 1 pt 0 353 mm If the mark is a circle the mark size corresponds to the diameter For other types of marks sim ilar conventions apply Initial value 6 defines the input mode to line and area drawing commands and c
237. will be rebuilt in DYANA according to the standard ge Ms File Formats ometry obtained from the residue library file Therefore conformations may be significantly changed if the Cartesian coordinates do not imply exactly the bond lengths bond angles and chiralities of the standard ge ometry The same applies for Cartesian coordinates where the dihedral angle of the peptide bonds are not exactly in the or conformation as de fined in the sequence input file An example output Cartesian coordinate file from DYANA follows Structure from DYANA f 2 50927E 01 DYANA 1 5 sgi 22 11 96 Number of residues 11 Number of atoms 240 1N 29 ARG 1 3249 0 0000 0 0000 2 HN 29 ARG 1 8841 0 0000 0 8290 3 CA 29 ARG 20133 0 0000 1 2455 4 HA 29 ARG 1 8760 0 9759 1 6891 5 CB 29 ARG 3 94 45 0 1727 0 9878 6 HB2 29 ARG 3 8976 0 5374 0 2278 7 HB3 29 ARG 3 7640 1 1709 0 5948 8 OB 29 ARG 3 8308 0 3167 0 4113 9 CG 29 ARG 4 3767 0 0400 2 2713 10 HG2 29 ARG 3 8459 0 4008 3 1151 11 HG3 29 ARG 4 4684 1 1070 2 4747 12 QG 29 ARG 4 1572 0 3531 2 7949 13 CD 29 ARG 5 7688 0 5840 2 1550 14 HD2 29 ARG 5 7220 1 4866 1 5457 15 HD3 29 ARG 6 1258 0 8829 3 1405 16 OD 29 ARG 5 9239 1 1847 2 3432 17 NE 29 ARG 6 7092 0 3855 1 5500 18 HE 29 ARG 7 2613 0 9490 2 1646 19 CZ 29 ARG 6 8670 0 5557 0 2303 20 NH1 29 ARG 7 7438 1 4591 0 2287 21 HH11 29 ARG 7 8615 1 5862 1 2136 22 HH12 29 ARG 8 2804 2 0063 0
238. wing steps An optional comment i e text following a comment sign FP is dis carded The values of variables are substituted from right to left The command line is split into elements defined as sequences of non blank characters separated by blank characters The first ele ment becomes the command name and the following elements be come command parameters If the command name matches a user defined alias the alias is ex panded If the command name matches a built in command of INCLAN it is executed by the command interpreter itself Otherwise if the command name matches a user defined command it is executed by the command interpreter Otherwise if the command name matches a command of the pro gram unambiguously it is executed by the program Otherwise the command interpreter looks for a macro with the given command name and if it is found in the current macro search path executes it If no such macro is found an error occurs 39 INCLAN 40 Special characters The following characters have a special meaning for INCLAN To use them literally they usually must be preceded by a backslash Svariable substitutes the value of the variable in the command line Substitutions proceed from left to right If the value of the variable or function call starts and ends with single quotes i e if it is a FORTRAN 77 character string the delimiting single quotes are removed before in serting
239. written out This command can only be used with distance constraint files file Write a Cartesian coordinate output file in PDB format the format used by the Brookhaven Protein Data Bank This command can only be used with coordinate files Execute a macro i e a file containing COFIMA commands A macro specification is the file specification of the macro file excluding the ex tension cfm Repeat the last command that started with string The string must not 181 COFIMA E contain spaces 182 Installation The program is delivered as a tar file dyana 1 5 tar possibly gzip compressed dyana 1 5 tar gz or compressed dyana 1 5 tar Z To uncompress the approriate commands are gunzip dyana 1 5 tar gz or uncompress dyana 1 5 tar Z Unpacking with the command tar xf dyana 1 5 tar creates in the current directory a subdirectory called dyana 1 5 that contains all files of the program package The different subdirectories contain the following data dyana DYANA source files inclan INCLAN source and help files macro DYANA standard macros lib residue libraries help on line help files example example files used in the tutorial cofima COFIMA source help and macro files scripts installation scripts The program is configured for a particular computer system by the shell script configure using the UNIX command 183 Installation 184 configure options install directory that
240. x axis and increases counterclock wise This command can also be used in list mode where the parameters are vector expressions See section mark 1Y1 2 92 draws a rectangle with corners x4 Y1 X1 Y2 Kos Y1 X Y2 and sets the current clipping path to its border Subsequent drawing com mands will only draw within this rectangular area off resets the clipping path After this command graphics will no longer be confined to the rectangular area specified in a previous Clip command closes the current output plot file 63 INCLAN DB comment cross curve curve dot errorbar 64 text writes text as a comment into the output plot file See section mark X1 Y1 X2 Y2 X3 Y3 NAYA normal mode draws a B zier spline curve defined by the points x y The total number of points must be 3n 1 with integer n2 1 The resulting curve passes through the points 1 4 7 the other points guide the curve Four points define the shape of each segment of the curve The curve segment leaves x y along the direction of the straight line connecting a y with x y2 and reaches x4 y4 along the direc tion of the straight line connecting x3 y3 with x4 y4 The lengths of the lines connecting x y with x5 yo and x3 y3 with x4 y4 represent in a sense the velocity of the path at the endpoints The curve segment is always enc
241. y default the importance level is normal Exits from the program file variable close Reads one line from a file and assigns it to a variable If the file is not yet open it is opened and the first line is read If the file is already open the next line is read If the end of the file is reached the variable is set to EOF and the file is closed Optionally the file can be closed after read ing a line file Removes one or more disk files exits from the current macro and returns to the calling macro or if the macro was called interactively to interactive input Given interactively return exits from the program variable value or if the keyword set is omitted variable value assigns a value i e a string to a variable set i 456 j 2 i Short form of set assigns a string value k 2 i Short form of eval evaluates an expression set i j k i 456 J 2 1 k 458 set show sleep subroutine syntax A INCLAN variable Displays values of variables If no variable is specified all variables that have values different from NULL are displayed If the names of one or several variables are given the values of these variables are displayed variable Displays the values of all or selected global variables If no variable is specified all global variables that have values different from NULL are displayed If the nam
242. ystem UNIX command Executes a UNIX command by invoking a shell If no command is speci fied an interactive shell is started type macro 61 INCLAN unset var 62 displays the macro or user defined command with the given name Mac ros in the current path can be listed without giving a path otherwise the path has to be specified variable Removes one or more variables variable declares variables as local variables of the current macro In contrast to normal global variables local variables are only visible within the macro where they are declared and within macros that are called via that macro except when such a macro declares itself a local variable with the same name The var command must precede any other commands in a macro except the parameter command and cannot be used interac tively Graphics With INCLAN it is possible to produce graphical output in either Post script of FrameMaker MIF format Graphics is created with the built in command plot The plot command can either be invoked directly or plot subcommands can be combined with list data in graphics files that can be read with the plot file command A graphics file can contain one or several blocks of list data i e matrices of integer or real numbers in free format Each row line of a list data block must have the same number of entries The columns of a list data block form vectors ca

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