Identification of Protease Specificity by Combining Proteome-Derived Peptide Libraries and Quantitative Proteomics.

ABSTRACT: We present protease specificity profiling based on quantitative proteomics in combination with proteome-derived peptide libraries. Peptide libraries are generated by endoproteolytic digestion of proteomes without chemical modification of primary amines before exposure to a protease under investigation. After incubation with a test protease, treated and control libraries are differentially isotope-labeled using cost-effective reductive dimethylation. Upon analysis by liquid chromatography-tandem mass spectrometry, cleavage products of the test protease appear as semi-specific peptides that are enriched for the corresponding isotope label. We validate our workflow with two proteases with well-characterized specificity profiles: trypsin and caspase-3. We provide the first specificity profile

SUBMITTER: Biniossek ML 

PROVIDER: S-EPMC4937521 | biostudies-literature | 2016 Jul

REPOSITORIES: biostudies-literature