Protease cleavage site fingerprinting by label-free in-gel degradomics reveals pH-dependent specificity switch of legumain.

ABSTRACT: Determination of protease specificity is of crucial importance for understanding protease function. We have developed the first gel-based label-free proteomic approach (DIPPS-direct in-gel profiling of protease specificity) that enables quick and reliable determination of protease cleavage specificities under large variety of experimental conditions. The methodology is based on in-gel digestion of the gel-separated proteome with the studied protease, enrichment of cleaved peptides by gel extraction, and subsequent mass spectrometry analysis combined with a length-limited unspecific database search. We applied the methodology to profile ten proteases ranging from highly specific (trypsin, endoproteinase GluC, caspase-7, and legumain) to broadly specific (matrix-metalloproteinase-3, thermoly

SUBMITTER: Vidmar R 

PROVIDER: S-EPMC5556264 | biostudies-literature | 2017 Aug

REPOSITORIES: biostudies-literature