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  1. out m e m b r a n e in EMBO Workshop, Cape Town, 2014

  2. 1. Tools-> Sequence -> Multialign Viewer 2. Choose “PF03595_seed.txt” 3. Select Aligned FASTA 4. Structure -> Render by Conservation EMBO Workshop, Cape Town, 2014

  3. out out out out EMBO Workshop, Cape Town, 2014

  4. 1. Actions-> Ribbon -> hide out out out out EMBO Workshop, Cape Town, 2014

  5. Chen et al. Nature 467 (2010) EMBO Workshop, Cape Town, 2014

  6. Chen et al. Nature 467 (2010) EMBO Workshop, Cape Town, 2014

  7. Chen et al. Nature 467 (2010) EMBO Workshop, Cape Town, 2014

  8. H. influenzaeprotein structure Functional hypothesis via homology to SLAC1 Identification potential functional residuesusing sequence conservation across the family and structural knowledge Suggested experiments to test functional hypothesis Thomine and Barbier-BrygooNature 467:1058-59 (2010) TUM, January 2013

  9. Anion channels exfoliative toxins malate uptake transporter sulphite efflux pump Chen et al. Nature 467 (2010) EMBO Workshop, Cape Town, 2014

  10. Example (using homology for protein annotation) EMBO Workshop, Cape Town, 2014

  11. >protein MESRSSPRLECSGAISAHCSLHLPDSSDFQLIFVFLVEMGFHHVGQAGLELLISSDLPTS ASQSAGITDMKLSMKNNIINTQQSFVTMPNVIVPDIEKEIRRMENGACSSFSEDDDSAST SEESENENPHARGSFSYKSLRKGGPSQREQYLPGAIALFNVNNSSNKDQEPEEKKKKKKE KKSKSDDKNENKNDPEKKKKKKDKEKKKKEEKSKDKKEEEKKEVVVIDPSGNTYYNWLFC ITLPVMYNWTMVIARACFDELQSDYLEYWLILDYVSDIVYLIDMFVRTRTGYLEQGLLVK EELKLINKYKSNLQFKLDVLSLIPTDLLYFKLGWNYPEIRLNRLLRFSRMFEFFQRTETR TNYPNIFRISNLVMYIVIIIHWNACVFYSISKAIGFGNDTWVYPDINDPEFGRLARKYVY SLYWSTLTLTTIGETPPPVRDSEYVFVVVDFLIGVLIFATIVGNIGSMISNMNAARAEFQ ARIDAIKQYMHFRNVSKDMEKRVIKWFDYLWTNKKTVDEKEVLKYLPDKLRAEIAINVHL DTLKKVRIFADCEAGLLVELVLKLQPQVYSPGDYICKKGDIGREMYIIKEGKLAVVADDG VTQFVVLSDGSYFGEISILNIKGSKAGNRRTANIKSIGYSDLFCLSKDDLMEALTEYPDA KTMLEEKGKQILMKDGLLDLNIANAGSDPKDLEEKVTRMEGSVDLLQTRFARILAEYESM QQKLKQRLTKVEKFLKPLIDTEFSSIEGPGAESGPIDST EMBO Workshop, Cape Town, 2014

  12. What is the function of our protein? EMBO Workshop, Cape Town, 2014

  13. Colour Scheme: N’ C’ 1MBN EMBO Workshop, Cape Town, 2014

  14. Colour Scheme: 1HW2 N’ C’ EMBO Workshop, Cape Town, 2014

  15. Colour Scheme: 1HW2 N’ C’ EMBO Workshop, Cape Town, 2014

  16. 1FOK 1HW2 Colour Scheme: N’ C’ EMBO Workshop, Cape Town, 2014

  17. 1FOK 1HW2 Colour Scheme: N’ C’ EMBO Workshop, Cape Town, 2014

  18. Z-score = 4.0 %id = 8% RMSD = 2.7Å EMBO Workshop, Cape Town, 2014

  19. 1FOK 1HW2 Colour Scheme: N’ C’ EMBO Workshop, Cape Town, 2014

  20. 1FOK 1HW2 Restriction endonuclease ? EMBO Workshop, Cape Town, 2014

  21. 1FOK 1HW2 Restriction endonuclease ? EMBO Workshop, Cape Town, 2014

  22. 1FOK 1HW2 Restriction endonuclease Transcription factor EMBO Workshop, Cape Town, 2014

  23. Definition (Wikipedia): A protein domain is a conserved part of a given protein sequence and structure that can evolve, function, and exist independently of the rest of the protein chain. Each domain forms a compact three-dimensional structure and often can be independently stable and folded. EMBO Workshop, Cape Town, 2014

  24. Also homologous domains can be functionally diverse Restriction endonuclease Transcription factor Human methionine aminopeptidase2 Vogel et al. Curr. Opin. Struct. Biol. 14 (2004) EMBO Workshop, Cape Town, 2014

  25. What is the function of our protein? EMBO Workshop, Cape Town, 2014

  26. >protein MKLSMKNNIINTQQSFVTMPNVIVPDIEKEIRRMENGACSSFSEDDDSASTSEESENENP HARGSFSYKSLRKGGPSQREQYLPGAIALFNVNNSSNKDQEPEEKKKKKKEKKSKSDDKN ENKNDPEKKKKKKDKEKKKKEEKSKDKKEEEKKEVVVIDPSGNTYYNWLFCITLPVMYNW TMVIARACFDELQSDYLEYWLILDYVSDIVYLIDMFVRTRTGYLEQGLLVKEELKLINKY KSNLQFKLDVLSLIPTDLLYFKLGWNYPEIRLNRLLRFSRMFEFFQRTETRTNYPNIFRI SNLVMYIVIIIHWNACVFYSISKAIGFGNDTWVYPDINDPEFGRLARKYVYSLYWSTLTL TTIGETPPPVRDSEYVFVVVDFLIGVLIFATIVGNIGSMISNMNAARAEFQARIDAIKQY MHFRNVSKDMEKRVIKWFDYLWTNKKTVDEKEVLKYLPDKLRAEIAINVHLDTLKKVRIF ADCEAGLLVELVLKLQPQVYSPGDYICKKGDIGREMYIIKEGKLAVVADDGVTQFVVLSD GSYFGEISILNIKGSKAGNRRTANIKSIGYSDLFCLSKDDLMEALTEYPDAKTMLEEKGK QILMKDGLLDLNIANAGSDPKDLEEKVTRMEGSVDLLQTRFARILAEYESMQQKLKQRLT KVEKFLKPLIDTEFSSIEGPGAESGPIDST EMBO Workshop, Cape Town, 2014

  27. >sp|P29973|CNGA1_HUMAN cGMP-gated cation channel alpha-1 OS=Homo sapiens GN=CNGA1 PE=1 SV=3 MKLSMKNNIINTQQSFVTMPNVIVPDIEKEIRRMENGACSSFSEDDDSASTSEESENENP HARGSFSYKSLRKGGPSQREQYLPGAIALFNVNNSSNKDQEPEEKKKKKKEKKSKSDDKN ENKNDPEKKKKKKDKEKKKKEEKSKDKKEEEKKEVVVIDPSGNTYYNWLFCITLPVMYNW TMVIARACFDELQSDYLEYWLILDYVSDIVYLIDMFVRTRTGYLEQGLLVKEELKLINKY KSNLQFKLDVLSLIPTDLLYFKLGWNYPEIRLNRLLRFSRMFEFFQRTETRTNYPNIFRI SNLVMYIVIIIHWNACVFYSISKAIGFGNDTWVYPDINDPEFGRLARKYVYSLYWSTLTL TTIGETPPPVRDSEYVFVVVDFLIGVLIFATIVGNIGSMISNMNAARAEFQARIDAIKQY MHFRNVSKDMEKRVIKWFDYLWTNKKTVDEKEVLKYLPDKLRAEIAINVHLDTLKKVRIF ADCEAGLLVELVLKLQPQVYSPGDYICKKGDIGREMYIIKEGKLAVVADDGVTQFVVLSD GSYFGEISILNIKGSKAGNRRTANIKSIGYSDLFCLSKDDLMEALTEYPDAKTMLEEKGK QILMKDGLLDLNIANAGSDPKDLEEKVTRMEGSVDLLQTRFARILAEYESMQQKLKQRLT KVEKFLKPLIDTEFSSIEGPGAESGPIDST EMBO Workshop, Cape Town, 2014

  28. Exercise (Find the structural domains) EMBO Workshop, Cape Town, 2014

  29. 2. File -> Open “2DHH_A.pdb” 1. Presets -> Interactive 1 (ribbons) 2. Presets -> Publication 1 (silhouette, rounded ribbons) 3. Actions -> Ribbon -> show 2DHH_A EMBO Workshop, Cape Town, 2014

  30. PA14 3ABZ-truncated EMBO Workshop, Cape Town, 2014

  31. Domain classification EMBO Workshop, Cape Town, 2014

  32. EMBO Workshop, Cape Town, 2014

  33. Class Class Architecture Fold Topology Superfamily Superfamily homology Family Family EMBO Workshop, Cape Town, 2014

  34. http://www.cathdb.info EMBO Workshop, Cape Town, 2014

  35. http://www.cathdb.info EMBO Workshop, Cape Town, 2014

  36. EMBO Workshop, Cape Town, 2014

  37. Protein family databases EMBO Workshop, Cape Town, 2014

  38. Building families Choose target Build MSA of representative members Build family profile MANUAL CURATION Use model to search sequence space for more members Annotate EMBO Workshop, Cape Town, 2014

  39. Profile Hidden Markov Models - Encapsulate diversity seq1 ACG-LD SCG--E NCGGFD TCG-WQ seq2 seq3 seq4 Profile-HMM slides courtesy of Rob Finn (EMBL-EBI) EMBO Workshop, Cape Town, 2014

  40. M1 Profile Hidden Markov Models - Encapsulate diversity seq1 ACG-LD SCG--E NCGGFD TCG-WQ seq2 seq3 seq4 N T A S Profile-HMM slides courtesy of Rob Finn (EMBL-EBI) EMBO Workshop, Cape Town, 2014

  41. M1 Profile Hidden Markov Models - Encapsulate diversity seq1 ACG-LD SCG--E NCGGFD TCG-WQ seq2 seq3 seq4 N T A S • C • M2 Profile-HMM slides courtesy of Rob Finn (EMBL-EBI) EMBO Workshop, Cape Town, 2014

  42. M1 Profile Hidden Markov Models - Encapsulate diversity seq1 ACG-LD SCG--E NCGGFD TCG-WQ seq2 seq3 seq4 N T A S • C G • M2 • M3 Profile-HMM slides courtesy of Rob Finn (EMBL-EBI) EMBO Workshop, Cape Town, 2014

  43. M1 Profile Hidden Markov Models - Encapsulate diversity seq1 ACG-LD SCG--E NCGgFD TCG-WQ seq2 seq3 seq4 N T A S • C G • M2 • M3 Profile-HMM slides courtesy of Rob Finn (EMBL-EBI) EMBO Workshop, Cape Town, 2014

  44. M1 Profile Hidden Markov Models - Encapsulate diversity seq1 ACG-LD SCG--E NCGgFD TCG-WQ seq2 seq3 seq4 W F L Y N T A S • C G • M2 • M4 • M3 Profile-HMM slides courtesy of Rob Finn (EMBL-EBI) EMBO Workshop, Cape Town, 2014

  45. M1 Profile Hidden Markov Models - Encapsulate diversity seq1 ACG-LD SCG--E NCGgFD TCG-WQ seq2 seq3 seq4 W F L Y N T A S D E Q • C G • M2 • M4 • M3 • M5 Profile-HMM slides courtesy of Rob Finn (EMBL-EBI) EMBO Workshop, Cape Town, 2014

  46. M1 Profile Hidden Markov Models - Encapsulate diversity seq1 ACG-LD SCG--E NCGgFD TCG-WQ seq2 seq3 seq4 W F L Y N T A S 123-45 D E Q • C G • M2 • M4 • M3 • M5 Profile-HMM slides courtesy of Rob Finn (EMBL-EBI) EMBO Workshop, Cape Town, 2014

  47. M1 Profile Hidden Markov Models - Encapsulate diversity seq1 ACG-LD SCG--E NCGgFD TCG-WQ seq2 seq3 seq4 W F L Y N T A S 123-45 D E Q • C G • M2 • M4 • M3 • M5 • I3 Profile-HMM slides courtesy of Rob Finn (EMBL-EBI) EMBO Workshop, Cape Town, 2014

  48. M1 Profile Hidden Markov Models - Encapsulate diversity seq1 ACG-LD SCG--E NCGgFD TCG-WQ seq2 seq3 seq4 W F L Y N T A S 123-45 D E Q • C G • M2 • M4 • M3 • M5 • D4 • I3 Profile-HMM slides courtesy of Rob Finn (EMBL-EBI) EMBO Workshop, Cape Town, 2014

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