Wiley Bioinformatics For Dummies, 2nd Edition
Contents
1. z N I So NH2 COOH 2 o So f z lt So N N I N 2 2 NH2 COOH So The protein molecule itself is made when a free NH group links chemically with a COOH group forming the peptide bond CO NH Figure 1 2 shows a schematic picture of the resulting chain As a result of this chaining process your protein molecule is going to be left with an unused NH at one end and an unused COOH at the other end These extremities are called respectively the N terminus and C terminus of the protein chain This is important to know because scientific convention in books databases and so on defines the sequence of a protein or of a protein fragment as the succession of its constituent amino acids listed in order from the N terminus to the C terminus The sequence of our short demo protein is then MAVLD Met Ala Val Leu Asp Methionine Alanine Valine Leucine Aspartic Working with protein 3 D structures The precise succession of a protein s constituent amino acids is what defines a given protein molecule This ribbon of amino acids however is not what NH2 CO NH CO NH CO NH CO NH COOH Chapter 1 Finding Out What Bioinformatics Can Do for You 15 MBER we amp gives the protein its biological properties for ins2. 1 Finding the best alignment between two or more proteins Chapters 8 and 9 1 Finding evolutionary relationships between proteins drawing proteins family trees Chapters 7 9 11 and 13 Analyzing DNA Sequences During the 1950s while scientists such as Kendrew and Perutz were still struggling to determine the first 3 D structures of proteins other biologists had already acquired a lot of indirect evidence via extremely clever genetics experiments that deoxyribonucleic acid DNA the stuff that makes up our genes was also a large macromolecule It was a long chainlike molecule twisted into a double helix and each link in the chain was a pairing of two out of four constituents called nucleotides A nucleotide is made up of one phos phate group linked to a pentose sugar which is itself linked to one of 4 types of nitrogenous organic bases symbolized by the four letters A C G and T However molecular biologists had to wait until much later the 1970s to be more precise before they could determine the sequence of DNA molecules and get direct access to the sequences of gene nucleotides This was a revolution earning A Sanger his second Nobel Prize because the small DNA sequence alphabet 4 nucleotides as compared to 20 amino acids allowed a much simpler and faster reading and quickly lent itself to complete automation Currently the worldwide rate of determining DNA sequences is faster by orders o
3. 16 Part I Getting Started in Bioinformatics De Figure 1 3 Example of protein 3 D structure schematic Protein bioinformatics covered in this book The study of protein sequences can get pretty complicated so compli cated in fact that it would take a pretty thick book to cover all aspects of the field We d like to take a more selective approach by focusing on those aspects of protein sequences where bioinformatic analyses can be most useful The following list gives you a look at some topics where such an analysis is particularly relevant to protein sequences and also tells which chapters of this book cover those topics in greater detail Retrieving protein sequences from databases Chapters 2 3 and 4 Y Computing amino acid composition molecular weight isoelectric point and other parameters Chapter 6 Computing how hydrophobic or hydrophilic a protein is predicting anti genic sites locating membrane spanning segments Chapter 6 Predicting elements of secondary structure Chapters 6 and 11 Predicting the domain organization of proteins Chapters 6 7 9 and 11 Visualizing protein structures in 3 D Chapter 11 Predicting a protein s 3 D structure from its sequence Chapter 11 Chapter 1 Finding Out What Bioinformatics Can Do for You 7 1 Finding all proteins that share a similar sequence Chapter 7 Classifying proteins into families Chapters 7 8 and 9
4. bases only a small percentage correspond to molecules that have actually been isolated by somebody or experimented upon That s because determining 24 Part I Getting Started in Bioinformatics the sequence of a protein is much more difficult than sequencing DNA but all the proteins that a given organism whether microbe or human being can synthesize are encoded in the DNA sequence of its genome Thus the smart shortcut that molecular biologists have been using is to read protein sequences directly at the information source in the DNA sequence This way we can pretend to know the amino acid sequence of a protein that has never been isolated in a test tube Turning DNA into proteins The genetic code When you know a DNA sequence you can translate it into the corresponding protein sequence by using the genetic code the very same way the cell itself generates a protein sequence The genetic code is universal with some exceptions otherwise life would be too simple and it is nature s solution to the problem of how one uniquely relates a 4 nucleotide sequence A T G C to a suite of 20 amino acids we re using symbols rather than actual chem icals to do the same Understanding how the cell does this was one of the most brilliant achievements of the biologists of the 1960s Yet the final answer can be contained in a miraculously small table as shown in Figure 1 9 Have a look but feel free to indulge in awed sile
5. mentary strands of a complete DNA molecule By complementarity we mean that a thymine T on one strand is always facing an adenine A and vice versa and guanine G is always facing a cytosine C These couples A T and G C although not linked by a chemical bond have a strict one to one reciprocal relationship When you know the sequence of nucleotides along one strand you can automatically deduce the sequence on the other one This amazing property and not the stylish helical structure is the Rosetta Stone that explains everything about DNA 20 Part I Getting Started in Bioinformatics MBER K amp Ce Figure 1 7 How two comple mentary strands can be read the same way i sequences For instance when living organisms reproduce each of their genes must be duplicated In order to do this nature doesn t go about it the way a photocopier would by making an exact copy Rather nature sepa rates the DNA strands and makes two complementary ones thanks to the magical two sided structure of DNA molecules This double strand structure of DNA makes the definition of a DNA sequence ambiguous Even with our convention of reading the nucleotides from the 5 end toward the 3 end you may decide to write down the bottom or the top sequence Convince yourself that they re both equally valid sequences by turning this book upside down Thus at each location a DNA molecule corre sponds to
6. Guanine Purine U Uracil Pyrimidine N Any nucleotide Purine or Pyrimidine continued 22 Part I Getting Started in Bioinformatics PC Figure 1 8 How RNA turns itself into a double stranded structure SSS Table 1 2 continued 1 Letter Code Nucleotide Base Name Category R AorG Purine Y CorU Pyrimidine ene None gap Some programs automatically handle the U instead of T conversion and many don t even distinguish between the two classes of nucleic acids So don t be surprised if a database entry displays RNA sequences such as mes senger RNA with a T instead of a U In fact like proteins RNA sequences are encoded in the DNA For this reason people have adopted the habit of work ing with the sequences of the RNA genes written in DNA rather than with RNA sequences RNA structures Playing with sticky strands Even though RNA molecules consist of single strands of nucleotides their natural urge for pairing with complementary sequences is still there Think of each such single strand as a free floating piece of Scotch tape You know that it won t take long for that tape to become a messy ball until no sticky part remains exposed This is exactly what happens to the single stranded RNA molecule more or less for the sake of poetic license although Figure 1 8 shows more precisely how the stickiness works C7 OS Now you understand why we insisted on the notion of strand compleme
7. word bioinformatics If you re good at what you do you ll want to know what all the fuss is about This chapter then is for you Instead of a formal definition that would take hours to cover all the ins and outs of the topic the best way to get a quick feel for what bioinformatics or swimming for that matter is all about is to jump right into the water that s what we do next Go ahead and get your feet wet with some basic mole cular biology concepts and the relevant questions intimately connected with such concepts that all together define bioinformatics Analyzing Protein Sequences If you eat steak you re intimately acquainted with proteins Your taste buds know them intimately anyway even if your rational mind was too busy with dinner to master the concept For you non steak lovers out there you ll be pleased to know that proteins abound in fish and vegetables too Moreover all these proteins are made up of the same basic building blocks called amino acids Amino acids are already quite complex organic molecules made of carbon hydrogen oxygen nitrogen and sulfur atoms So the overall recipe for a protein the one your rational mind will appreciate even if your taste buds won t is something like C1200H24000600N3005100 Chapter 1 Finding Out What Bioinformatics Can Do for You The early days of biochemistry were devoted to finding out a better way to represent proteins preferably in
8. Chapter 1 Finding Out What Bioinformatics Can Do for You In This Chapter Defining bioinformatics Understanding the links between modern biology genomics and bioinformatics Determining which biological questions bioinformatics can help you answer quickly Organic chemistry is the chemistry of carbon compounds Biochemistry is the study of carbon compounds that crawl Mike Adam looks like biologists are colonizing the dictionary with all these bio words we have bio chemistry bio metrics bio physics bio technology bio hazards and even bio terrorism Now what s up with the new entry in the bio sweepstakes bio informatics What Is Bioinformatics In today s world computers are as likely to be used by biologists as by any other highly trained professionals bankers or flight controllers for example Many of the tasks performed by such professionals are common to most of us We all tend to write lots of memos and send lots of e mails many of us use spreadsheets and we all store immense amounts of never to be seen again data in complicated file systems However besides these general tasks biologists also use computers to address problems that are very specific to biologists which are of no interest to bankers or flight controllers These specialized tasks taken together make up the field of bioinformatics More specifically we can define bioinformatics as the computational branch of molecular biology 10 P
9. Val V GCG Ala A GAG Glu E GGG Gly G T A G 26 Part I Getting Started in Bioinformatics Because of the triplet based genetic code a given DNA interval on a given strand can theoretically be translated in three different ways basically three perspectives that are known in the field as reading frames Because the DNA can be used from both strands a total of six possible reading frames are possi ble for translating a DNA sequence into proteins With very few exceptions found in exotic viruses only one of these six frames is used for any given DNA coding region An interval of DNA sequence that remains free of STOP the translation of TAA TGA or TAG is called an open reading frame ORF Additional complications arise from the fact that some DNA sequences are not encoding proteins at all and that higher organisms have large pieces of noncoding DNA inserted within their genes A large part of bioinformatics is devoted to the development of methods to locate protein coding regions in DNA sequences to delineate precisely where genes start and end or where they are interrupted by the noncoding intervals called introns DNA RNA bioinformatics covered in this book Need a road map to the bioinformatic analyses that are relevant to DNA RNA sequences covered in this book Here it is Retrieving DNA sequences from databases Chapters 2 and 3 Computing nucleotide compositions Chapter 5 1 Identifying r
10. ally biochemists discovered that these amino acids are linked together as a chain and that the true identity of a protein is derived not only from its composition but also from the precise order of its constituent amino acids The first amino acid sequence of a protein insulin was determined in 1951 The actual recipe for human insulin from which all its biological prop erties derive is the following chain of 110 residues insulin MALWMRLLPLLALLALWGPDPAAAFVNQHLCGSHLVEALYLVCGERG FFYTPKTRREAEDLQVGQVELGGGPGAGSLQPLALEGSLOKRGIVEQCCTSICSLY QLENYCN Now more than 50 years later analyzing protein sequences like these remains a central topic of bioinformatics in all laboratories throughout the world Check out Chapters 2 4 and 6 through 11 to quickly figure out how to analyze your protein sequence and become a member of the club A brief history of sequence analysis Besides earning Alfred Sanger his first Nobel Prize the sequencing of insulin inaugurated the modern era of molecular and structural biology Traditionally a soft science that is more tolerant of fuzzy reasoning and hand waving ambiguity than chemistry or physics biology got a taste of its first fundamental dataset molecular sequences In the early 1960s known protein sequences accumulated slowly perhaps a blessing in disguise given that the computers capable of analyzing them hadn t been developed In this pre computer era from our present perspective an
11. art I Getting Started in Bioinformatics Time for a little bit of history Before the era of bioinformatics only two ways of performing biological experiments were available within a living organism so called in vivo or in an artificial environment so called in vitro from the Latin in glass Taking the analogy further we can say that bioinformatics is in fact in silico biology from the silicon chips on which microprocessors are built This new way of doing biology has certainly become very trendy but don t think that trendy translates into lightweight or flash in the pan Bioinformatics goes way beyond trendy it s at the center of the most recent developments in biology such as the deciphering of the human genome another buzzword system biology trying to look at the global picture new biotechnologies new legal and forensic techniques as well as the personalized medicine of the future Because of the centrality of bioinformatics to cutting edge developments in molecular biology people from many different fields have been stumbling across the term in a variety of different contexts If you re a biology medical or computer science student a professional in the pharmaceutical industry a lawyer or a policeman worrying about DNA testing a consumer concerned about GMOs Genetically Modified Organisms or even a NASDAQ investor interested in start up companies you ll already have come across the
12. biological roles For instance most DNA cut ting enzymes so called restriction enzymes have palindromic target sequences Other palindromic sequences serve as binding sites where regula tory proteins stick so they can turn genes on and off Palindromic sequences also have a strong influence on the 3 D structure of DNA molecules And not just DNA See the next section for more on palindromic sequences in RNA Looking for exact or approximate palindromes in DNA sequences is a classic bioinformatic exercise Analyzing RNA Sequences DNA deoxyribonucleic acid is the most dignified member of the nucleic acid family of macromolecules Its sole and only task is to ensure forever the conservation of the genetic information for its organism It is thus very stable and resistant and lies well protected in the nucleus of each cell Ribonucleic acid RNA is a much more active member of the nucleic acid family it s syn thesized and degraded constantly as it makes copies of genes available to the cell factory In the context of bioinformatics there are only two important differences between RNA and DNA 1 RNA differs from DNA by one nucleotide 1 RNA comes as a single strand not a helix The one letter IUPAC codes for RNA sequences are shown in Table 1 2 Table 1 2 Most Common Letters Used for RNA Nucleotide Sequences 1 Letter Code Nucleotide Base Name Category A Adenine Purine C Cytosine Pyrimidine G
13. es would be encoded by similar sequences of amino acids The function of a protein turned out to be a direct consequence of its 3 D structure shape The resulting logical linkage SEQUENCE STRUCTURE FUNCTION was established and is now a central concept of molecular biology and bioinformatics Playing with protein structure models on a computer screen is of course much easier than carrying around a thousand piece 3 D plastic puzzle As a consequence an increasing proportion of the bioinformatics pie is now devoted to the development of cyber tools to navigate between sequences and 3 D structures This specialized area is called structural bioinformatics Thanks to many free resources on the Internet it is not difficult to display some beautiful protein pictures on your own computer and start playing with them as in video games We show you how to do that in Chapter 11 Before you get a chance to read that chapter Figure 1 3 gives you an idea of what a 400 amino acid typical protein 3 D schematic structure looks like when you don t have a color monitor and can t make it move and turn Don t forget Protein molecules even in their wonderful complexity are still pretty small The one in Figure 1 3 would fit in a box whose sides mea sure 70 1 000 000 millimeters There are thousands of different proteins in a single bacterium each of them in thousands of copies more than enough evidence that Life Is Not Simple
14. estriction sites Chapter 5 Designing polymerase chain reaction PCR primers Chapter 5 Identifying open reading frames ORFs Chapter 5 Predicting elements of DNA RNA secondary structure Chapter 12 1 Finding repeats Chapter 5 1 Computing the optimal alignment between two or more DNA sequences Chapters 7 8 and 9 1 Finding polymorphic sites in genes single nucleotide polymorphisms SNPs Chapter 3 vy Assembling sequence fragments Chapter 5 Working with Entire Genomes The first truly efficient technique to sequence DNA was discovered in 1977 In 1995 the first sequence of an entire genome from the microbe Hemophilus influenzae was determined Between these two dates DNA Chapter 1 Finding Out What Bioinformatics Can Do for You sequencing technologies improved steadily but such technologies still tended to concentrate on mining individual genes for information During this period biologists were mostly sequencing DNA fragments that were a few thousand nucleotides in length simply because they were interested in spe cific genes that they had started working on years before Most of the bioin formatics tools available today were created during that period They include All basic sequence alignment programs 1 Phylogenetic and classification methods 1 Various display tools adapted to relatively small sequence objects such as protein sequences no more than a few thousand characters long Genom
15. f magnitude than the rate of protein sequencing Reading DNA sequences the right way As was the case for the 20 amino acids found in proteins the 4 nucleotides making DNA have different bodies but all have the same pair of hooks 5 phosphoryl and 3 hydroxyl pronounced five prime and three prime by reference to their positions in the deoxyribose sugar molecule which is part of the nucleotide chaining device Figure 1 4 shows what free individual nucleotides look like Forming a bond between the 5 and 3 positions of the constituent nucleotides then makes the DNA molecule Figure 1 5 shows a schematic representation of the resulting DNA strand 18 Part I Getting Started in Bioinformatics a Figure 1 4 The four nucleotides making DNA PC Figure 1 5 Chained nucleotides constituting a DNA strand a MBER we amp 5 P 3 0H 5 P 3 0H 5 P 3 OH 5 P 3 0H 5 P 3 5 3 5 3 5 3 5 3 0H After the nucleotides are linked the resulting DNA strand exhibits an unused phosphoryl group PO at the 5 end and an unused hydroxyl group OH at the 3 end These extremities are respectively called the 5 terminus and the 3 terminus of the DNA strand A DNA sequence is always defined in books databases articles and pro grams as the succession of its constituent nucleotides listed from the 5 to 3 terminus that is end The sequence of the short DNA strand shown in Figure 1 5 is then TGACT Thy
16. ics Getting all the genes at once The determination of the first complete genome sequence terminated the gene by gene routine and initiated the era of genomics the genetic mapping physical mapping and sequencing of entire genomes As a consequence the DNA sequences we have to work with now are much longer close toa million bp in length for microbes and up to several billion bp in length for animals and humans This revolution called for the design of new bioinformatic tools and databases capable to store query analyze and display these huge objects in a user friendly manner Chapters 3 5 and 7 present some of the questions that biologists address at the genome scale and show the relevant bioinformatic tools in action In contrast to the early days of the gene by gene approach DNA sequences are now often obtained along with the presumed protein sequences derived from those DNA sequences without any prior knowledge of what is actually there In essence genes are both sequenced and discovered at the same time This development prompted the emergence of an entirely new branch of bioinformatics devoted to the parsing of large DNA sequences into their components genes transcription units protein coding regions regulatory elements and so forth This first pass is then followed by a longer phase of genome annotation where the biological functions of these various elements are more or less tentatively predicted Part IV of this book
17. k you ll find more in depth coverage of specific topics some of them bristling with scary mouth filling terms related to genome bioinformatics Finding which genomes are available Chapter 3 Analyzing sequences in relation to specific genomes Chapters 3 and 7 Displaying genomes Chapter 3 Parsing a microbial genome sequence ORFing Chapter 5 Parsing a eukaryotic genome sequence GenScan Chapter 5 Finding orthologous and paralogous genes Chapter 3 Finding repeats Chapter 5
18. l Pyrrolysine U Sec Selenocysteine Z Gln or Glu Glutamine or glutamic acid X Xaa Any residue The B and Z codes which are now becoming obsolete indicated how hard it was to distin guish between Asp and Asn or Glu and Gin in the early days of protein sequence determina tion In contrast the J code shows how difficult it is to distinguish between Ile and Leu using mass spectrometry the latest sequencing tech nique The Pyl and Sec exotic amino acids are No corresponding residue gap specified by the UAG Pyl and UGA Sec stop codons read ina specific context The X code is still very much used as a placeholder letter when you don t know the amino acid at a given position in the sequence Alignment programs use to denote positions apparently missing from the sequence Reading protein sequences from N to C The twenty amino acid molecules found in proteins have different bodies their characteristic residues listed in Table 1 1 but all have the same pair of hooks NH and COOH These groups of atoms are used to form the so called peptidic bonds between the successive residues in the sequence Figure 1 1 shows free individual amino acids floating about displaying their hooks for all to see 13 14 Part I Getting Started in Bioinformatics SE Figure 1 1 Free amino acids floating around PC LEE Figure 1 2 Amino acids chained together to constitute a protein molecule aay
19. mine Guanine Adenine Cytosine Thymine The two sides of a DNA sequence In the same laboratory where Kendrew and Perutz were trying to figure out the first 3 D structure of a protein Watson and Crick elucidated in 1953 the famous double helical structure of the DNA molecule These days every body has a mental picture of this famous spiral staircase molecule the ele gance of the DNA double helix probably helped make it the most popular notion to come out of molecular biology But what made this discovery so important earning one more Nobel Prize for molecular biology was not the helical shape but the discovery that the DNA molecule consists of two complementary strands shown in Figure 1 6 Chapter 1 Finding Out What Bioinformatics Can Do for You 9 NBER A The IUPAC code for DNA sequences The following table lists the one letter codes of Pure and Applied Chemistry are defined for all IUPAC codes used to work with DNA sequences possible two and three way ambiguities The Official IUPAC codes fromthe International Union table shows only the ones most frequently used Most Common Letters Used for DNA Nucleotide Sequences 1 Letter Code Nucleotide Name Category A Adenine Purine C Cytosine Pyrimidine G Guanine Purine T Thymine Pyrimidine N Any nucleotide any base n a R AorG Purine Y CorT Pyrimidine co None gap 7 S 5 gt 3 Figure 1 6 The two comple
20. n these sticky sequences Go directly to Chapter 12 if your main interest is in RNA bioinformatics More on nucleic acid nomenclature Don t panic if you get the impression that books courses and the technical literature all use many different words and abbreviations to designate the building blocks of nucleic acids That s actually true for example you ll find base base pair nucleoside and nucleotide but note These different names designate slightly different chemical entities and those differences are irrelevant for us just now So far we ve used the term nucleotide abbreviated nt as in a 400 nt long sequence This way of labeling a sequence refers to the length of the DNA or RNA molecules in terms of the number of positions they have available for nucleotides For instance the sequence in Figure 1 5 is 5 nt long Notice that we say number of positions rather than number of nucleotides A 400 nt long DNA molecule has 400 positions for nucleotides but it actually contains twice that many 800 because every position contains a pair of nucleotides To make this clearer DNA sequence sizes are often given in base pairs abbreviated bp Thus the DNA sequence in Figure 1 5 is 5 bp long Larger units such as kb 1000 bp or Mb mega bp are also used DNA Coding Regions Pretending to Work with Protein Sequences Of the hundreds of thousands of protein sequences found in current data
21. nce as you enter the most sacred monument of modern biology Here s how to use the table shown in Figure 1 9 From a given starting point in your DNA sequence start reading the sequence 3 nucleotides one triplef at a time Then consult the genetic code table to read which amino acid corre sponds to the current triplet technically referred to as codons For instance the following DNA or messenger RNA sequence is decoded as follows 1 Read the DNA sequence ATGGAAGTATTTAAAGCGCCACCTATTGGGATATAAG 2 Decompose it into successive triplets ATG GAA GTA TTT AAA GCG CCA CCT ATT GGG ATA TAAG 3 Translate each triplet into the corresponding amino acid MEVFKAPPIGISTOP If your DNA sequence is correctly listed in the 5 to 3 orientation you gener ate the protein sequence in the conventional N to C terminus as well This approach has an advantage You don t have to think about these orientation details ever again Thus if you know where a protein coding region starts in a DNA sequence your computer can pretend to be a cell and generate the corresponding amino acid sequence This simple computer translation exercise is at the E Figure 1 9 The universal genetic code Chapter 1 Finding Out What Bioinformatics Can Do for You 25 origin of most of the so called protein sequences that you can find in data bases Many sequence analysis programs acknowledge this fact by offering on the fly translation so you can process DNA se
22. ntarity refer to Figure 1 6 Single stranded RNA molecules pair different regions of their sequences to form stable double helical structures admittedly less regular than but quite similar to the double helical structure of DNA Once synthesized each RNA molecule quickly adopts a compact fold trying to pair as many nucleotides as possible while keeping the chain not only flexi ble but true to its own geometry Hairpin shapes as shown in Figure 1 8 are Chapter 1 Finding Out What Bioinformatics Can Do for You 23 the basic elements of RNA secondary structure they re made up of loops the unpaired C U in Figure 1 8 and stems the paired regions Just for fun verify for yourself that a palindromic RNA sequence results in a perfect hairpin with no loop While attempting to pair as many nucleotides as possible the RNA chain folds in space resulting in a specific 3 D structure that s dictated by its sequences As with proteins the linear sequence of the building blocks dictates the final 3 D shape The biological function of RNA molecules derives from their 3 D shapes or from their sequence complemen tarity with specific genes Computing predicting the final fold of an RNA molecule from its sequence is a challenging problem that drove many historical developments in bioinfor matics The recent discovery that small RNA molecules can switch off the activity of a number of genes is what triggered a renewed interest i
23. presents you with some of these most advanced techniques Figure 1 10 representing the whole genome of the bacterium Rickettsia conorii illustrates this new level of complexity This circular DNA molecule is 1 3 million bp long on the small side for a bacterium Each little rectangle in the two most external circles of features one circle per strand corresponds to a protein coding gene in the circular genome Each rectangle corresponds to approximately 1000 bp Nobody knew which genes or which proteins were in that bacterium before the sequencing started Almost everything we know now about this bacterium and many others we can describe as fairly inaccessible such as those thriving on the ocean floor near volcanic vents at 100 C has been derived from bioinformatic analyses 27 28 Part Getting Started in Bioinformatics 300 000 400 000 200 000 aN va yyy Wiw t UAT N WY 1 Mig 4 syn V tay f HI auum o l Y R KN yw Mt y My Minn 100 000 S N h Wy ay Ge 600 000 R conorii 1 268 755 bp 650 000 700 000 Figure 1 10 A Represen tation of a 800 000 bacterial genome n 900 006 1 000 000 1 200 000 Myf iy Y Mngt i w Htp Min wt 7 Many MN W 1 OUN ra ys My n N 1 f m tama ws WS 1 100 000 A Ming Th Y Genome bioinformatics covered in this book The following list lets you know where in this boo
24. quences as virtual protein sequences with a simple mouse click More with coding DNA sequences Using the example in the first paragraphs of the section DNA Coding Regions Pretending to Work with Protein Sequences you can see that the resulting protein sequence depends entirely on the way you converted your DNA sequence into triplets before using the genetic code For instance using the second position as starting point leads to 1 ATGGAAGTATTTAAAGCGCCACCTATTGGGATATAAG 2 A TGG AAG TAT TTA AAG CGC CAC CTA TTG GGA TAT AAG 3 MK EOX R B b ly E iK Beginning with the third position GGA AGT again leads to an entirely different translation Table of Standard Genetic Code T A G TTT Phe F TCT Ser S TAT Tyr Y TGT Cys TTC Phe F TCC Ser S TAC Tyr Y TGC Cys C TTA Leu L TCA Ser S TAA Stop TGA Stop TTG Leu L TCG Ser S TAG Stop TGG Trp W CTT Leu L CCT Pro P CAT His H CGT Arg R c CTC Leu L CCC Pro P CAC His H CGC Arg R CTA Leu L CCA Pro P CAA Gln Q CGA Arg R CTG Leu L CCG Pro P CAG Gln Q CGG Arg R ATT Ile I ACT Thr T AAT Asn N AGT Ser S ATC Ile Q ACC Thr T AAC Asn N AGC Ser S ATA Ile I ACA Thr T AAA Lys K AGA Arg R ATG Met M ACG Thr T AAG Lys K AGG Arg R GTT Val V GCT Ala A GAT Asp D GGT Gly G GTC Val V GCC Ala A GAC Asp D GGC Gly G GTA Val V GCA Ala A GAA Glu E GGA Gly G GTG
25. tance its ability to digest sugar or to become part of a muscle fiber those come from the three dimensional 3 D shape that the ribbon adopts in its environment A protein molecule once made is not a chainlike highly flexible object think like a section of chain link fence rather it s more like a compact well bundled ball of string The final 3 D shape of the protein molecule is uniquely dictated by its sequence because some amino acid types for instance hydrophobic residues L V I have no desire whatsoever to be at the surface interacting with the surrounding water while others for instance hydrophilic residues D S K are actively looking for such an opportunity The protein chain is also affected by other influences such as the electric charges carried by some of the amino acids or their capacity to fit with their immediate neighbors The first 3 D structure of a protein was determined in 1958 by Drs Kendrew and Perutz using the complicated technique of X ray crystallography Not for the faint of heart Don t grapple with how it works unless you want to turn professional Besides winning one more Nobel Prize for the nascent field of molecular biology this feat made the doctors realize that proteins have precise and specific shapes encoded in the sequence of amino acids Hence they pre dicted that proteins with similar sequences would fold into similar shapes and conversely that proteins with similar structur
26. terms of a formula that would explain their biological or even nutritional properties Biochemists realized over time that proteins were huge molecules macromolecules made up of large numbers of amino acids typically from 100 to 500 picked out from a selec tion of 20 flavors with names such as alanine glycine tyrosine glutamine and so on Table 1 1 gives you the list of these 20 building blocks with their full names three letter codes and one letter codes the JUPAC code after the International Union of Pure and Applied Chemistry committee that designed it Table 1 1 The 20 Amino Acids and Their Official Codes 1 Letter Code 3 Letter Code Name 1 A Ala Alanine 2 R Arg Arginine 3 N Asn Asparagine 4 D Asp Aspartic acid 5 C Cys Cysteine 6 Q Gin Glutamine 7 E Glu Glutamic acid 8 G Gly Glycine 9 H His Histidine 10 l lle Isoleucine 11 L Leu Leucine 12 K Lys Lysine 13 M Met Methionine 14 F Phe Phenylalanine 15 P Pro Proline 16 S Ser Serine 17 T Thr Threonine 18 W Trp Tryptophan 19 Y Tyr Tyrosine 20 V Val Valine 2 Part I Getting Started in Bioinformatics Biochemists then recognized that a given type of protein such as insulin or myoglobin always contains precisely the same number of total amino acids generically called residues in the same proportion Thus a better formula for a protein looks like this insulin 30 glycines 44 alanines 5 tyrosines 14 glutamines Fin
27. two totally different sequences related by this reverse and complement operation This isn t complicated simply keep it in mind every time you work with DNA sequences Fortunately most database mining programs such as BLAST know about this property and take both strands into account when reporting their results But some programs don t bother and only analyze the sequence you gave them In cases where both strands matter always make sure that a complete analysis has been performed We discuss these details further in Chapters 3 5 and 7 Palindromes in DNA sequences Newcomers to DNA sequence analysis are usually confused by the notion of reverse complementary sequences However in due time you ll be able to recognize right away that the two sequences ATGCTGATCTTGGCCATCAATG and CATTGATGGCCAAGATCAGCAT correspond to facing strands of the same DNA molecule One fascinating property of DNA complementarity is the fact that regions of DNA may correspond to sequences that are identical when read from the two complementary strands Figure 1 7 helps illustrate this magic trick 5 gt 3 Chapter 1 Finding Out What Bioinformatics Can Do for You 2 Such sequences are called palindromes after the term for a phrase or sen tence that reads the same in both directions such as Madam I m Adam or A man a plan a canal Panama Palindromic sequences aren t merely a curiosity they play important
28. yway sequences were assembled analyzed and compared by manually writing them on pieces of paper taping them side by side on laboratory walls and or moving them around for optimal alignment now called pattern matching As soon as the early computers became available as big as locomotives and just as fast and with 8K of RAMD the first computational biologists started to enter these manual algorithms into the memory banks This practice was brand new nobody before them had to manipulate and analyze molecular sequences as texts Most methods had to be invented from scratch and in the process a new area of research the analysis of protein sequences using computers was generated This was the genesis of bioinformatics Chapter 1 Finding Out What Bioinformatics Can Do for You Seven additional amino acid codes When you work with databases or analysis pro grams you re likely to have some unusual let ters popping up now and then in your protein sequences These letters are either used to designate exotic amino acids or are used to denote various levels of ambiguity that is a total lack of information about certain posi tions in the sequence We ve listed these par ticular letters in the following table Seven Codes for Ambiguity or Exceptional Amino Acids 1 Letter Code 3 Letter Code Meaning B Asn or Asp Asparagine or aspartic acid J Xle Isoleucine or leucine O letter Py
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