title: Improving peptide identification sensitivity in shotgun proteomics by stratification of search space.
abstract: Because of its high specificity, trypsin is the enzyme of choice in shotgun proteomics. Nonetheless, several publications do report the identification of semitryptic and nontryptic peptides. Many of these peptides are thought to be signaling peptides or to have formed during sample preparation. It is known that only a small fraction of tandem mass spectra from a trypsin-digested protein mixture can be confidently matched to tryptic peptides. If other possibilities such as post-translational modifications and single-amino acid polymorphisms are ignored, this suggests that many unidentified spectra originate from semitryptic and nontryptic peptides. To include them in database searches, however, may not improve overall peptide identification because of the possible sensitivity reduction from search space expansion. To circumvent this issue for E-value-based search methods, we have designed a scheme that categorizes qualified peptides (i.e., peptides whose differences in molecular weight from the parent ion are within a specified error tolerance) into three tiers: tryptic, semitryptic, and nontryptic. This classification allows peptides that belong to different tiers to have different Bonferroni correction factors. Our results show that this scheme can significantly improve retrieval performance compared to those of search strategies that assign equal Bonferroni correction factors to all qualified peptides.
authors.affiliation: National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, Maryland 20894, United States.
authors.forename: Gelio
authors.initials: G
authors.lastname: Alves
authors.forename: Yi-Kuo
authors.initials: YK
authors.lastname: Yu
mesh_terms: D000465:Algorithms; D000818:Animals; D006801:Humans; D015233:Models, Statistical; D058977:Molecular Sequence Annotation / Q000706:statistics & numerical data*; D010446:Peptide Fragments / Q000302:isolation & purification*; D059748:Proteolysis; D040901:Proteomics; D012680:Sensitivity and Specificity; D020539:Sequence Analysis, Protein / Q000706:statistics & numerical data*; D053719:Tandem Mass Spectrometry; D014357:Trypsin / Q000737:chemistry
chemical_list: D010446:Peptide Fragments; D014357:Trypsin