蛋白质组学基本原理幻灯片.ppt

* Characteristics of various search engines Search engine Key features Scoring method Mascot MS and MS/MS modes combined Automated searches are available Probability based MS-Fit/MS-Tag MS or MS/MS modes Automated searches not available MOWSE in MS mode match count in MS/MS mode PeptIdent http://www.expasy.ch/ tools/peptident.html MS mode only Automated searches not available Genetic algorithm based on the characteristics of TOF MS spectra ProFound/Sonars MS or MS/MS modes Automated searches available Bayesian approach SEQUEST MS/MS mode only Automated searches available Cross-correlation method The search parameters of database search engines(Mascot) mass accuracy MS mode mass accuracy MS mode amino acid modifications organism info database name mass of precursors reporting features enzyme specificity charge state of precursors Sequence databases and information sources for protein identification Protein sequences from protein sequencing Translated from full length cDNA sequences Translated from incomplete cDNA sequences (EST) Predicted from raw genomic sequences Source Reliability Volume of data Examples high high medium low low medium high high SWISSPROT SWISSPROT, TrEMBL, nrNCBI nrNCBI dbEST Peptide Mass Fingerprinting:An Example with a Real Peptide 14 peptides resulted from tryptic digestion of human hemoglobin alpha chain； One peptide VGAHAGEYGAEALEA with monoisotopic mass of 1528.7348 Da； Searching in the SWISS-PROT database (mouse and human proteins) yields two matches: IGGHGAEYGAEALER mouse 1529.7348 VGAHAGEYGAEALER human 1529.7348 Accurate protein identification requires multiple peptide matches。 Peptide Mass Fingerprinting:Limitations Imperfect mass data on peptides due to limited mass accuracy of instrument； Too many signals to be accurately identified； False-positive matches due to chance alone, rather than actual identify； Incomplete protein digestion by protease； Misrepresention of modified proteins in the database； Protein to be identifi