UCSF Chimera - GroEL Visualization
Contents
1. The full GroEL can be seen by looking at the crystal unit cell Use menu entry Tools Multiscale Multiscale Models and near the bottom of the Multiscale dialog choose Crystal unit cell for the Multimer type and press the Make Models button no Ea r The Multiscale dialog makes copies of PDB chains using symmetry matrices It displays the copies as low resolution surfaces Each of the 7 chains of a ring is given a different color You will see 4 GroEL barrels Each barrel consists of two rings of PDB model 1OEL stacked to form a cylinder M Coloring monomers Select chains All With loaded atoms Clear You can select individual surfaces using Ctrl left ET i m mouse button and change their color with the E Color Well button in the middle of the Selected chains show Hide Hide allsyles Multiscale dialog If you select a surface and then _ srw mi _sie ates press the Copies button near the top of the a a Multiscale dialog it will extend the selection to Calor atoms _rivoons to maton surfaces all copies of that chain gone erection Ea Delete chains Models from molecules and matrices Showing ribbons Multimer Crystal unit cell MTRIX SMTRY CRYST1 a l Make models Delete selected models Close Help Jj Press the With loaded atoms button in the selection pane at the top of the Multiscale dialog _ and towards the middle of the d2. 080 map In the File Open File Select Actions Tools Favorites Help dialog you should change the File Type to All guess type Initially it is set to only show Protein Data Bank files pdb About this density map This is a single particle reconstruction of GroEL determined by Steve Ludtke using EMAN software GroEL is a cylindrical 14 mer that refolds misfolded proteins in its interior cavity This density map and others are available from the EM database at the Macromolecular Structure Database no selection This GroEL map has 11 5 angstrom resolution Another map determined by Steve having 6 angstrom resolution is also available We ll use the lower resolution map because it is easier to see the main features Moving the map Dragging the mouse in the graphics window lets you rotate translate and zoom in on the map e Left mouse button rotation e Middle mouse button translation e Right mouse button zooming On a Mac with a one button mouse hold down the Option key to get middle mouse button behaviour or the Command key to get right mouse button behaviour Volume Viewer dialog v ROE Ax Controls for display of volume Panels _j Data W Display J Region _s Options data ie density maps are in the pata ema1o80map size 100 100 100 step 111 Volume Viewer dialog that was sece v mesh w solid 7 Ctrl click on histogram to add or delete thresholds displayed when you opened the pecs
3. Model Panel dialog menu entry Favorites Model Panel by clicking the checkbutton in the Shown column next to emd_1080 map Display the density map as a mesh using the Volume Viewer dialog menu entry Tools Volumes Volume Viewer Rotate with the mouse so you are looking directly down the axis of GroEL cylinder with the 1OEL model near you Now switch off the Active m Model Panel checkbutton next to emd_1080 map in the Model Panel dialog This in the column before the Shown button and each model has a separate checkbutton Active means that the model can be moved with the mouse Now use the middle mouse button to drag the PDB model so that it is centered with the density map frequently used Configure Close Help Rotating about the axis perpendicular to the screen Once the PDB model is centered with the density map drag with the left mouse button near the edge of the graphics window to rotate the PDB model to match the shape of the density map When you do a mouse rotation at the edge of the window this rotates about the axis perpendicular to the screen Using the mouse more towards the middle rotates the model about axes parallel to the screen as if you are grabbing the model The boundary between these mouse rotation zones is not shown Turn on the Active button on the Model Panel for emd_1080 map so you can rotate both models at once and check the alignment from different view points Undoing acc
4. UCSF Chimera GroEL Visualization The purpose of this exercise is to get familiar with Chimera features for density map display and display of Protein Data Bank PDB models Outline of what follows e Density map basics e Working with atomic models e Hand aligning PDB models in density maps e Image saving Followed by more advanced features e Tricks for better images e Density map slicing and coloring e Using symmetry matrices e Displaying b factor as ribbon thickness e Aligning protein domains And miscellaneous topics e Toolbar buttons e Chimera manual Help buttons e Session saving e More tutorials Starting Chimera On Linux run the executable chimera in the bin directory of your Chimera installation If Chimera is installed in usr local chimera then you run usr local chimera bin chimera from a shell On Windows start Chimera by double clicking the Chimera icon in the directory called bin in your Chimera installation If Chimera is installed in Program Files then the executable will be found in directory Program Files Chimera bin On Mac start the Apple X server found in Applications Utilities X11 Then double click the Chimera application to start The X server is not installed in Mac OS 10 3 by default and if you do not have it you can download it from Apple It is necessary for running Chimera on the Mac Open GroEL density map Use the Chimera File Open dialog to MESSET TF open emd_1
5. an control how shiny the models appear using menu entry Tools Viewing Parameters Shininess Control The shininess parameter effects the size of specular highlights Specular highlights are the white lighting spots on the model The brightness parameter just effects the brightness of the highlights not the whole surface v UCSF Chimera x File Select Actions Tools Slicing surfaces Favorites Help The Tools Graphics Per Model Clipping menu entry lets you slice through surfaces and protein models Set the model to emd_1080 map and turn on the Enable clipping switch in the dialog that is shown Then rotate the density map to see the cut The cut surface is visually confusing because you see inside of the surface Use the Tools Graphics Surface Capping menu entry and switch on Cap surfaces at clip planes This will cap the hole where the surface was cut up arrow to increase se no selection Moving the clip plane To move the cut plane also called the clipping plane switch on the Adjust clipping with mouse switch in the Per Model Clipping dialog Then drag the clip plane with the middle mouse button or rotate it with the right mouse button Turn off the Adjust clipping with mouse switch to restore the mouse buttons their normal behaviour of translating models and zooming E Fer Clipping Ax Model emd_1080 map 0 W Enable clipping Orient plane perpendicular to line of sight wit
6. ave default dialog settings button at the bottom of the Options panel Chimera settings are saved in a hidden directory called chimera in your home directory in a file called preferences Griw 7 1 Camera Effects Rotation Depth cueing Chimera dims parts of the scene that are further from you to give a sense of depth This feature can be turned on and off using menu entry Tools Viewing Parameters Effects The dimming occurs between the near and far clip planes The clip planes can be adjusted using menu entry Tools Resolution high Viewing Parameters Side View The Side View window displays the open Eee aa models as seen from the side Two vertical yellow lines indicate the front and back clip planes The left line is the front clip plane If you drag the front clip plane yellow vertical line towards the models with the mouse you will see the brightness in the main graphics window increase provided depth cueing is turned on If you move the front clip plane so it intersects the models you will see the surface cut in the main graphics window The small yellow box at the left in the side view is your eye position You can drag this left or right to zoom in and out Lighting and Shininess The Tools Viewing Parameters Lighting menu entry lets you move the 2 Chimera lights The key light provides the main illumination of your models while the fill light is intended to fill in dark areas You c
7. display now First hide the density map saraelecon using menu entry Favorites Model Panel and switching off the checkbutton in the Shown column for emd_1080 map v Dee 5 clipping ID Name Active Shown close 0 emd_1080 map v z iv color by 55 inl focus frequently used aa Configure Close Help Showing a ribbon representation To show a ribbon display of the model use menu entry Actions Ribbon show To hide the original all atom display use menu entry Actions Atom Bonds hide Zooming in shows a flat ribbon To make the ribbon have an oval cross section use Actions Ribbon round Coloring the ribbon secondary structure To color the alpha helices beta sheets and VES 777777 turns of the ribbon using separate colors l Select Actions Tools Favorites Help click on the 1OEL pdb line in the Model Panel dialog and then press the Color by SS button in the righthand column of that dialog The SS is short for secondary structure Press the OK button on the color secondary structure dialog to use the default colors Coloring individual chains To color one of the 7 proteins in this model differently select a single residue using the mouse 3997 atoms 3876 bonds e Ctrl left mouse button select objects The selected residue will be highlighted with a green outline It may be hard to see since it is small Pre
8. ee aie ge cepa raat Ee 5 wll ry can be shown again with menu Range 1 34 2 37 Level 1 67 Color i entry Tools Volumes Volume snonnes p Bini Viewer Transparency Factor 0 am Show Unshow Center Orient L Open Remove Close Help J The center histogram shows the distribution of data values in the density map The range of data values 1 34 to 2 37 is shown below the histogram The white vertical bar on the histogram shows the contouring threshold Drag this bar with the mouse to change the contour level The numeric value of the contour level is shown below the histogram in the entry field Contour levels can be typed in instead of being set with the mouse Contour level Color Clicking on the white square with the wide raised MENEM 5 x border below the histogram shows a Color Editor dialog The square button called a Color Color name femman Well is used in many Chimera dialogs to choose Color space RGB 4 Opacity colors and shows the current color in the center r 1o00 Change the density map color by clicking on the j fo color bars in the Color Editor dialog You can ae close the dialog when you are done Mesh display style Above the histogram is a radio button where you can choose mesh display style This shows the surface as a mesh made up of triangles You may need to zoom in to see the small triangles Data size and subsampling Near the top o
9. f the dialog the data size 100 by 100 by 100 is shown The Step menu allows you to subsample the data For example a step of 2 2 2 means to use every other density value along each of the 3 axes Subsampling is useful with large maps to speed up moving and recontouring Outline box The outline box shows the bounds of the data set You can hide the outline box by clicking the Options checkbutton on the right side of the top line of the Volume Viewer dialog This displays a panel of options and the first one Show outline box can be switched off Clicking the Options checkbutton again hides the long panel of options Brighter image To make the density map display brighter use menu entry Tools Viewing Parameters Effects and switch off Depth cueing in the Viewing dialog Depth cueing makes objects appear dimmer the further they are away from you This will be discussed more in the section on tricks for better images It is a good idea to close Chimera dialogs when you are done so your whole screen does not become covered with dialogs Working with atomic models We now look at an atomic model of ana GroEL determined by x ray Help crystallography and how this can be aligned with the density map Use File Open to open Protein Data Bank model 1OEL pdb It is in the same directory as the density map The atomic model of one half of GroEL will be shown shifted to one side of the density map We ll look at methods of atomic model
10. ferences Choose the Tools category The entries in this list of tools are exactly those in the Tools menu in the same order Check buttons in the second column of the scrollable list of tools to put a button that invokes that menu entry on the Toolbar at left side of the main window For example check Inspectors Model Panel and Viewing Parameters Side View If you want these buttons to appear every time you start Chimera press the Save button at the bottom of the Preferences dialog Chimera manual Help buttons Pressing the Help button at the bottom of any Chimera dialog will show the appropriate section of the Chimera manual included with the Chimera distribution in your web browser Almost all Chimera dialogs have these help buttons The user manual documentation is very thorough and worth looking at Also look at the Help menu on the main window Session saving You can save your Chimera session with File Save Session As Later you can restart Chimera and use File Restore Session to redisplay your models This will save the PDB models Multiscale surfaces density map display settings colorings background color and many other attributes It does not save all aspects of the session The surface clipping capping and coloring shown before are not saved in our current version but may be in future releases The session is saved as a Python file py file suffix It saves a copy of the PDB data but does not copy the densi
11. h center of rotation at center of view Flip plane 160 degrees Align plane with emd_1080 map 0 cp plane Cap style solid mesh Mesh subdivision factor 1 0 Cap to clip plane distance 0 001 Close Help NESE Capping 5 W Cap surfaces at clip planes _j Use cap color W Adjust clipping with mouse as below Clip Motion Assignments Translation Rotation button 2 nd button Fd Close Help i Surface coloring The density map values vary across the clipped capped surface The variation can be shown by coloring the cap using Tools Volumes Surface Color Change the Color scheme in the Surface Color dialog to be 5 color The 3 color color scheme is intended for display of electrostatic potentials To only color the cap switch on Color surface caps only Then press the Color button at the bottom of the dialog When you move the clip plane the coloring will be automatically updated v pU GE 50x z Eile Select Actions Tools Favorites Help Y EES x Color surface emd_1080 map ad Using volume data emd_1080 map at Color scheme 5 color x 5 E N t ar co par oo nN pos Color outside volume ri W Color surface caps only Stop recoloring surface Color Close Help up arrow to increase selection atoms re no selection Clip perpendicular to GroEL cylinder To clip the GroEL density perpend
12. h or height entry fields and press the Enter key You only need to enter one of the dimensions When you press Enter the other will be calculated to maintain the aspect ratio and the image will be captured To change the pixels per inch press the Image Setup button If you want an image exactly the same number of pixels as the Chimera window on the screen press the Get Pixels button This fills in the exact size in pixels of the Chimera graphics window White background When making images sometimes you want the background to be white Do this using menu entries Actions Color background followed by Actions Color white Tricks for better images Smooth surfaces Smoothing off Smoothing on Zooming in on the surface you can see facets from the triangles use surface display style To reduce these you can turn on the surface smoothing Click the Options checkbutton at the top of the Volume Viewer dialog and check the Surface smoothing iterations checkbutton the last entry in the options panel The list of options is so long it may go off the bottom of your screen You may need to hide the Display panel by unchecking the Display checkbutton at the top of the Volume Viewer dialog to see the full list of options You can leave this option on its only serious drawback is that it can slow down recontouring the surface To have make your changed Volume Viewer settings the defaults for future Chimera sessions press the S
13. h to the Worms tab and panutes ot resaues 2 eee press Apply Render Select Attribute average gt bfactor 4 Ctr click on histogram to add or delete thresholds The ribbon thickness now corresponds to average residue b factor value The low b factor face of 1OEL lie at the interface between the two 7 mer rings and the high b factor face is at the end of the barrel Value 147 087 Worm raius Colors Radii Worms Worm style smooth Restrict OK Apply to current selection if any If you had a chain selected only that chain shows the thickness variation If you clear the selection Ctrl click on empty space or menu entry Select Clear Selection and press Apply again all chains will show b factor OK Apply Close Help Adjusting worm thickness The Render by attribute dialog shows a histogram of the b factor values Vertical bars on the histogram mark B factor values which have a specified thickness For b factor values between the bars the ribbon radius is linearly interpolated using the values defined for the bars Clicking on a histogram bar shows the corresponding ribbon thickness in the Worm radius entry field below The bars can be moved and radii changed Unlike the Volume Viewer dialog where moving the histogram bar causes an immediately updated display here you need to press Apply to see the effect of your change Aligning protein domains GroEL adopts d
14. ialog use the Style Show button which pops up a menu and select Ribbon This shows your original copy of 1OEL Deleting extra GroEL barrels Delete the 3 extra copies of the GroEL barrel by selecting each barrel with the mouse and pressing the Delete Chains button in the Multiscale dialog You can select a barrel with the mouse using Ctrl left mouse button and dragging a box around it This selects everything in the rectangle you drag out highlighted in green To avoid selecting molecules you do not wish to delete it is best to drag a small box only a piece of each molecule need be in the box to be selected Even if a molecule is hidden from view behind another one it will be selected if it partially lies in the box you draw UCSF chimera 7 AE File Select Actions Tools Favorites Help Displaying b factor as ribbon thickness The LOEL PDB model is a crystal structure that has a B factor associated with each atom that indicates how well localized the atom is in the x ray density To display the B factor using ribbon thickness select the 1OEL pdb line in the Model Panel and press the Render by attribute button in the right column of that dialog It will take several seconds to scan the attributes of this large 27 000 atom structure v Render Select by Attribute 0x Choose Attributes of residues in the Render by Se attribute dialog choose Attribute to be average SETE gt b factor then switc
15. icular to the cylindrical axis first press the Orient button on at the bottom of the Volume Viewer dialog then press the Orient plane perpendicular to line of sight button on the Per Model Clipping dialog The Orient button on the volume dialog rotates the density map back to the original orientation when the file was first opened z axis perpendicular to screen After orienting the plane try moving it with the middle mouse button Accurate surface cap colors If you change the contouring threshold in the Volume Viewer dialog by dragging the marker on the histogram the colors on the capped surface will change by small amounts Ideally they should not change and the problem is that the volume data is not being sampled at enough points on the cut surface You can see the sampling by switching Cap style to mesh in the Surface Capping dialog The mesh switch in the Surface Capping dialog effects just the cap while the mesh switch in the Volume Viewer dialog applies to just the contour surface Increasing the Mesh subdivision factor in the Surface Capping dialog to 2 0 press Enter after changing the number for change to take effect will cause cap vertices to be more closely spaced by a factor of 2 Moving the contour threshold then produces very little artifactual color variation but updates are slower Crystal unit cell The 1OEL crystal structure has only one of the two 7 protein rings in its asymmetric unit GroEL has two such rings stacked
16. idental rotation If you accidentally rotate the PDB model so its axis is not parallel to the density map cylinder axis use menu entry Tools Utilities Undo Move to bring them back to having parallel cylinder axes Image saving File Save Image Screen shot jagged edges Use menu entry File Save Image and press the Save As button to save the graphics window image to a file After the image is captured a dialog appears to choose the file name and format Image capture directly from screen a Saveimage BEE m Image Size The Chimera dialogs will disappear and you will Unites nenes i C g A a PP y Image width 2 71473 see magnified pieces of the image flash by in the graphics window Chimera is capturing the image directly from the screen so no dialog should be placed on top of the Chimera window while the image is taken Image height 2 94977 Maintain current aspect ratio j hr Get Pixels Image Setup Image Information Supersampling Description H Using Chimera image saving will produce a higher quality image than taking a screen Image Credits Citing Chimera snapshot The image is captured at a size 3 times _save as cancer Her larger than requested and then averaged down to the correct size This is called supersampling It reduces jagged edges Large image sizes You can make images larger than the screen for publications and posters Type a new value into the dialog widt
17. ifferent conformations and our hand alignment of PDB model 1OEL and density map emd_1080 is only a first step in comparing this crystal and single particle EM data The GroEL monomer has 3 domains that can assume different orientations relative to one another A next step would be to computationally fit the individual protein domains into the EM map Chimera does not compute optimal positioning of PDB models or domains in density maps You would need to use other software to do that Here are a series of Chimera images showing the 3 domains of the GroEL monomer Two monomers in PDB model 1AON having identical amino acid sequence but different conformations are compared Several steps were used to produce these images that we will not describe here Some of the tools used to make these images are e Multiscale Models menu Tools Multiscale e Scale Bar menu Tools Utilities e Match Maker menu Tools Homology Multalign Viewer structure matching capability menu Tools Homology e Model Panel write PDB menu Tools Inspectors or Favorites Toolbar buttons You can put buttons for frequently use Tools on the left side of the main Chimera window using Favorites Preferences v eae Tools m Category File Select Actions Settings O Toolbar EHEHEHEH E KIL LLU Reset Restore Save Close Help The Preferences dialog has several categories of pre
18. ssing the up arrow key on the keyboard the one used for moving up one line in a text editor will extend the selection to the entire chain containing that residue Then use menu entry Actions Color cyan to color the selected chain Hiding a ribbon To select a chain by its PDB identifier use menu Select Chain which contains entries A B C D E F G corresponding to the 7 chains in PDB model 1OEL Choose any chain from the menu to select it To hide the selected chain use Actions Ribbon hide You use Ribbon hide instead of Atoms Bonds hide because the chain is displayed as a ribbon The hide menu entries for the two styles are separate because you can have both ribbon and atoms bonds showing at the same time Selecting more than one chain To select multiple chains select a residue from one chain with Ctrl left mouse button Then select a residue in another chain with Shift Ctrl left mouse button Adding the Shift key extends the existing selection instead of replacing it Then press the up arrow key to select both chains Slow rotation When an object is selected the model may not rotate smoothly because drawing the green outline around the object requires alot of computation To clear the selection click Ctrl left mouse on empty space or use menu entry Select Clear Selection Hand align PDB model into density Now we will move the PDB model to align it with the density map Redisplay the density map using the
19. ty map data For density maps it saves the path to the original file For large PDB models such as 1OEL 27 000 atoms saving and restoring the session can take a few minutes More tutorials Chimera has many tools not explored in this tutorial Most are for analyzing and comparing protein structures and sequences More tutorials can be found in the Chimera User s Guide using the Chimera Help Tutorials menu entry or possibly more recent ones on the Chimera web site http www cgl ucsf edu chimera in the User s Guide
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