Journal
PROTEOMICS
Volume 10, Issue 15, Pages 2833-2844Publisher
WILEY-V C H VERLAG GMBH
DOI: 10.1002/pmic.200900821
Keywords
Bioinformatics; Chymotrypsin; CNBr; Protease specificity; Proteolysis; V8 protease
Funding
- NIH [5R01RR016522-05, 1-P41-RR024851-01]
- Howard Hughes Medical Institute
- NIH Center of Proteomics Research Resource for Integrative Biology [RR018522]
- DOE [DE-AC05-76RLO 1830]
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Although trypsin remains the most commonly used protease in MS, other proteases may be employed for increasing peptide coverage or generating overlapping peptides. Knowledge of the accurate specificity rules of these proteases is helpful for database search tools to detect peptides, and becomes crucial when label-free MS is used to discover in vivo proteolytic cleavages. Since in vivo cleavages are inferred by subtracting digestion-induced cleavages from all observed cleavages, it is important to ensure that the specificity rule used to identify digestion-induced cleavages are broad enough to capture even minor cleavages produced in digestion, to avoid erroneously identifying them as in vivo cleavages. In this study, we describe MS-Proteolysis, a software tool for identifying putative sites of in vivo proteolytic cleavage using label-free MS. The tool is used in conjunction with digestion by trypsin and three other proteases, whose specificity rules are revised and extended before inferring proteolytic cleavages. Finally, we show that comparative analysis of multiple proteases can be used to detect putative in vivo proteolytic sites on a proteome-wide scale.
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