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Part II : Sequence Comparison Multiple Sequence Alignment

Part II : Sequence Comparison Multiple Sequence Alignment. By Zhiwei Cao Dept. of Biological Science National university of Singapore Email: dbsczw@nus.edu.sg. Pair-Wise Alignment : Two Sequences. Multiple sequence alignment -- MSA.

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Part II : Sequence Comparison Multiple Sequence Alignment

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  1. Part II : Sequence ComparisonMultiple Sequence Alignment By Zhiwei Cao Dept. of Biological Science National university of Singapore Email: dbsczw@nus.edu.sg

  2. Pair-Wise Alignment : Two Sequences Made by Cao Zhiwei

  3. Multiple sequence alignment -- MSA • The multiple sequence alignment problem is to simultaneously align more than two sequences. Made by Cao Zhiwei

  4. Multiple sequence alignment Made by Cao Zhiwei

  5. What is MSA: A Definition Residues 2D table Absolute and relative positions Sequences Made by Cao Zhiwei

  6. Why multiple sequence alignment 1. Determine whether a group of proteins are related 2. Show regions of conservation within a protein family  sequence pattern 3. Determine evolutionary history of gene families  phylogeny tree Made by Cao Zhiwei

  7. MSA: How to Align? Seq1 AGAC Seq2 AC Seq3 AG Seq1 AGAC Seq2 --AC Seq2 AC Seq3 AG Seq1 AGAC Seq3 AG-- Made by Cao Zhiwei

  8. MSA: Some Possible Alignments Made by Cao Zhiwei

  9. MSA History • Until 1987 multiple alignments constructed manually from pairwise alignments • Lipman et al. 1989 pairwise dynamic programming approach applied to multiple sequence alignment - MSA http://www.psc.edu/general/software/packages/msa/msa.html Made by Cao Zhiwei

  10. Commonly Used MSA Methods • Dynamic programming - extension of pairwise sequence alignment • Progressive sequence alignment - incorporates phylogenetic information to guide the alignment process • Iterative sequence alignment - correct for problems with progressive alignment by repeatedly realigning subgroups of sequence Made by Cao Zhiwei

  11. Progressive Method of MSA • Progressive alignment invented in ‘87 & ‘88 - Feng & Doolittle 1987, Higgins and Sharp 1988 • Based on phylogeny Made by Cao Zhiwei

  12. How MSA: Progressive method Scerevisiae [1] Celegans [2] 0.640 Drosophia [3] 0.634 0.327 Human [4] 0.630 0.408 0.420 Mouse [5] 0.619 0.405 0.469 0.289 1 - Do pairwise alignment of all sequences and calculate distance matrix [1] [2] [3] [4] 2 1 Made by Cao Zhiwei

  13. Human Mouse Dmel Cele Scer How MSA: Progressive method 2 - Create a guide tree based on this pairwise distance matrix Made by Cao Zhiwei

  14. How MSA: Progressive method 3 - Align progressively following guide tree • Start by aligning most closely related pairs of sequences • Gaps • At each step align two sequences or one to an existing subalignment Made by Cao Zhiwei

  15. Available programs for progressive MSA • CLUSTAL (Free package): • Higgins,D.G. and Sharp,P.M. (1988) CLUSTAL: a package for performing multiple sequence alignment on a microcomputer. Gene 73,237-244. • http://www.ebi.ac.uk/clustalw/ • http://clustalw.genome.ad.jp/ (origin 2) • PILEUP (part of GCG commercial package) • http://www.gcg.com • Others Made by Cao Zhiwei

  16. Example software---ClustalWhttp://clustalw.genome.ad.jp Made by Cao Zhiwei

  17. Example Software---ClustalW (Bioedit)http://www.mbio.ncsu.edu/BioEdit/bioedit.html Made by Cao Zhiwei

  18. Steps To Do ClustalW: Step 1: Prepare the sequences: • Retrieve sequences • General considerations: • The more the better • Exclude similar (>80%) sequences • Necessary modification Made by Cao Zhiwei

  19. Steps To Do ClustalW: Step 2: Input the sequences: • Put all sequnces into one file Copy and paste • Upload sequences one by one • Pay attention to sequence format Made by Cao Zhiwei

  20. Steps To Do ClustalW: Step 3: Set the parameters: • Default parameters for protein alignment General Setting Parameters: • Output Format: CLUSTALW • Pairwise Alignment: FAST/APPROXIMATE Made by Cao Zhiwei

  21. Example: SH2 domainfamily • SH2 domains function as regulatory modules of intracellular signalling cascades • V-Src Tyrosine Kinase Transforming Protein (Phosphotyrosine Recognition Domain Sh2) Complex With Phosphopeptide A (PDB code 1SHA): Made by Cao Zhiwei

  22. Input Sequences For ClustalW • >1SHA-A V-SRC Tyrosine kinase transforming protein (SH2 domain), from Rous sarcoma virus • >1A81-A Chain A, Tandem Sh2 Domain Of The Syk Kinase, from Homo sapiens • >1JWO-A Chain A, Sh2 Domain Of The Csk Homologous Kinase Chk, from Homo sapiens • >1BLJ Nmr Ensemble Of Blk Sh2 Domain, from Mus musculus (house mouse) Made by Cao Zhiwei

  23. Result 1 of ClustalW Made by Cao Zhiwei

  24. Result 2 of ClustalW Made by Cao Zhiwei

  25. Result 3 of ClustalW: N-J tree Made by Cao Zhiwei

  26. Interpret ClustalW results • Three characters are used in the results 2: • '*' indicates positions which have a single, fully conserved residue • ':'indicates that 'strongly' conserved groups • '.'indicates that 'weakerly' conserved groups Made by Cao Zhiwei

  27. Interpret ClustalW results • Insertion and deletion, gap Consensus ………………………QCGG………....G.....C …......C...........YSQC... Consensus sequenceSequence Pattern Made by Cao Zhiwei

  28. Notes on how to use ClustalW • Remove signal peptide before alignment, try to compare homologous portion • Sequence containing a repetitive element (such as a domain) • Heuristic algorithm: not guaranteed for perfect alignment Made by Cao Zhiwei

  29. Notes on how to use ClustalW • Mobilize your biological knowledge, check the alignment and recheck the alignment • Manually re-align your sequences if it’s bad Made by Cao Zhiwei

  30. Application of MSAExample: Drug discovery for SARSAnand et al., www.scienceexpress.org//10.1126/science.1085658, published May 13, 2003 • Coronaviruses are positive-stranded RNA viruses • Sequence structure function • Human coronavirus 229E: HCoV; • Porcine transmissible gastroenteritis virus: TGEV; • Mouse hepatitis virus: MHV; • Bovine coronavirus: BCoV; • SARS-associated coronavirus: SARS-CoV; • Avian infectious bronchitisvirus: IBV. Made by Cao Zhiwei

  31. Application of MSA Example: Drug Discovery for SARS Anand et al., www.scienceexpress.org//10.1126/science.1085658, published May 13, 2003 Made by Cao Zhiwei

  32. Summary • What is MSA • Why do MSA • How to do MSA • Available computational methods • ClustalW • Interpret results of ClustalW • Quality control • Application example of MSA: SARS drug discovery Made by Cao Zhiwei

  33. Phylogeny tree: evolutionary history Made by Cao Zhiwei

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