1. Protein analysis and proteomics Friday, 27 January 2006 Introduction to Bioinformatics DA McClellan david_mcclellan@byu.edu

2. protein [1] Protein families [4] Protein function [2] Physical properties [3] Protein localization Fig. 8.1 Page 224

3. Perspective 1: Protein families (domains and motifs) Page 225

4. Definitions Signature: a protein category such as a domain or motif Domain: a region of a protein that can adopt a 3D structure a characteristic fold or functional region a family (superfamily) is a group of proteins that share a domain examples: zinc finger domain immunoglobulin domain Motif (or fingerprint): a short, conserved region of a protein typically 10 to 20 contiguous amino acid residues Page 225

5. 15 most common domains (human) Zn finger, C2H2 type1093 proteins Immunoglobulin1032 EGF-like471 Zn-finger, RING458 Homeobox417 Pleckstrin-like405 RNA-binding region RNP-1400 SH3394 Calcium-binding EF-hand392 Fibronectin, type III300 PDZ/DHR/GLGF280 Small GTP-binding protein 261 BTB/POZ236 bHLH226 Cadherin226 Table 8-3 Page 227 Source: Integr8 program at www.ebi.ac.uk/proteome/

6. Definition of a domain According to InterPro at EBI ( http://www.ebi.ac.uk/interpro /): A domain is an independent structural unit, found alone or in conjunction with other domains or repeats. Domains are evolutionarily related. Tables 8-1,8-2 Page 226 According to SMART (http://smart.embl-heidelberg.de): A domain is a conserved structural entity with distinctive secondary structure content and a hydrophobic core. Homologous domains with common functions usually show sequence similarities.

7. Varieties of protein domains Fig. 8.2 Page 228 Extending along the length of a protein Occupying a subset of a protein sequence Occurring one or more times

8. Example of a protein with domains: Methyl CpG binding protein 2 (MeCP2) MBD Page 227 TRD The protein includes a methylated DNA binding domain (MBD) and a transcriptional repression domain (TRD). MeCP2 is a transcriptional repressor. Mutations in the gene encoding MeCP2 cause Rett Syndrome, a neurological disorder affecting girls primarily.

9. Fig. 8.3 Page 228 Result of an MeCP2 blastp search: A methyl-binding domain shared by several proteins

10. Are proteins that share only a domain homologous? Fig. 8.3 Page 228

11. ProDom entry for HIV-1 pol shows many related proteins Fig. 8.7 Page 231

12. Proteins can have both domains and patterns (motifs) Domain (aspartyl protease) Domain (reverse transcriptase) Pattern (several residues) Pattern (several residues) Fig. 8.7 Page 231

13. Fig. 8.8 Page 232

14. Definition of a motif A motif (or fingerprint) is a short, conserved region of a protein. Its size is often 10 to 20 amino acids. Simple motifs include transmembrane domains and phosphorylation sites. These do not imply homology when found in a group of proteins. PROSITE (www.expasy.org/prosite) is a dictionary of motifs (there are currently >1300 entries)(9/05). In PROSITE, a pattern is a qualitative motif description (a protein either matches a pattern, or not). In contrast, a profile is a quantitative motif description. We will encounter profiles in Pfam, ProDom, SMART, and other databases. Page 231-233

15. Perspective 2: Physical properties of proteins Page 233

16. Fig. 8.9 Page 234 Posttranslational modifications:

17. Fig. 8.11 Page 235

18. Fig. 8.11 Page 235

19. Fig. 8.12 Page 236

20. Fig. 8.13 Page 238

21. Fig. 8.13 Page 238

22. Fig. 8.13 Page 238

23. Syntaxin, SNAP-25 and VAMP are three proteins that interact via coiled-coil domains

24. Introduction to Perspectives 3 and 4: Gene Ontology (GO) Consortium Page 237

25. The Gene Ontology Consortium An ontology is a description of concepts. The GO Consortium compiles a dynamic, controlled vocabulary of terms related to gene products. There are three organizing principles: Molecular function Biological process Cellular compartment You can visit GO at http://www.geneontology.org. There is no centralized GO database. Instead, curators of organism-specific databases assign GO terms to gene products for each organism. Page 237

26. GO terms are assigned to Entrez Gene entries Fig. 8.14 Page 241

27. Fig. 8.14 Page 241

28. Fig. 8.14 Page 241

29. Fig. 8.14 Page 241

30. The Gene Ontology Consortium: Evidence Codes ICInferred by curator IDAInferred from direct assay IEAInferred from electronic annotation IEPInferred from expression pattern IGIInferred from genetic interaction IMPInferred from mutant phenotype IPIInferred from physical interaction ISSInferred from sequence or structural similarity NASNon-traceable author statement NDNo biological data TASTraceable author statement Table 8-7 Page 240

31. Perspective 3: Protein localization Page 242

32. protein Protein localization Page 242

33. Protein localization Proteins may be localized to intracellular compartments, cytosol, the plasma membrane, or they may be secreted. Many proteins shuttle between multiple compartments. A variety of algorithms predict localization, but this is essentially a cell biological question. Page 242

35. Fig. 8.15 Page 242 PSORT: searches for sorting signals that are characteristic of proteins localized to particular cellular compartments

36. Fig. 8.16 Page 244

37. Fig. 8.16 Page 244

38. Localization of 2,900 yeast proteins Michael Snyder and colleagues incorporated epitope tags into thousands of S. cerevisiae cDNAs, and systematically localized proteins (Kumar et al., 2002). See http://ygac.med.yale.edu for a database including 2,900 fluorescence micrographs. Page 243

39. Perspective 4: Protein function Page 243

40. Protein function Function refers to the role of a protein in the cell. We can consider protein function from a variety of perspectives. Page 243

41. 1. Biochemical function (molecular function) RBP binds retinol, could be a carrier Fig. 8.17 Page 245

42. 2. Functional assignment based on homology RBP could be a carrier too Other carrier proteins Fig. 8.17 Page 245

43. 3. Function based on structure RBP forms a calyx Fig. 8.17 Page 245

44. 4. Function based on ligand binding specificity RBP binds vitamin A Fig. 8.17 Page 245

45. 5. Function based on cellular process DNARNA RBP is abundant, soluble, secreted Fig. 8.17 Page 245

46. 6. Function based on biological process Analyze a gene knockout phenotype; RBP is essential for vision Fig. 8.17 Page 245

47. 7. Function based on “proteomics” or high throughput “functional genomics” High throughput analyses show... RBP levels elevated in renal failure RBP levels decreased in liver disease Fig. 8.17 Page 245

48. Functional assignment of enzymes: the EC (Enzyme Commission) system Oxidoreductases1,003 Transferases1,076 Hydrolases1,125 Lyases356 Isomerases156 Ligases126 Table 8-8 Page 246 Updated 9/04, 9/05

49. Functional assignment of proteins: Clusters of Orthologous Groups (COGs) Information storage and processing Cellular processes Metabolism Poorly characterized Table 8-9 Page 247 See Chapter 14 for COGs at NCBI