title: A bead-based cleavage method for large-scale identification of protease substrates.
abstract: Proteolysis is a major form of post translational modification which occurs when a protease cleaves peptide bonds in a target protein to modify its activity. Tracking protease substrates is indispensable for understanding its cellular functions. However, it is difficult to directly identify protease substrates because the end products of proteolysis, the cleaved protein fragments, must be identified among the pool of cellular proteins. Here we present a bead-based cleavage approach using immobilized proteome as the screening library to identify protease substrates. This method enables efficient separation of proteolyzed proteins from background protein mixture. Using caspase-3 as the model protease, we have identified 1159 high confident substrates, among which, strikingly, 43.9% of substrates undergo degradation during apoptosis. The huge number of substrates and positive support of in vivo evidence indicate that the BBC method is a powerful tool for protease substrates identification.
authors.affiliation: Key Laboratory of Separation Sciences for Analytical Chemistry, National Chromatographic R&A Center, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China.|Institute of Cancer Stem Cell, Dalian Medical University, Dalian 116044, China.
authors.forename: Chunli
authors.initials: C
authors.lastname: Wang
authors.affiliation: Key Laboratory of Separation Sciences for Analytical Chemistry, National Chromatographic R&A Center, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China.
authors.forename: Mingliang
authors.initials: M
authors.lastname: Ye
authors.affiliation: Key Laboratory of Separation Sciences for Analytical Chemistry, National Chromatographic R&A Center, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China.
authors.forename: Xiaoluan
authors.initials: X
authors.lastname: Wei
authors.affiliation: Key Laboratory of Separation Sciences for Analytical Chemistry, National Chromatographic R&A Center, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China.
authors.forename: Yangyang
authors.initials: Y
authors.lastname: Bian
authors.affiliation: Key Laboratory of Separation Sciences for Analytical Chemistry, National Chromatographic R&A Center, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China.
authors.forename: Kai
authors.initials: K
authors.lastname: Cheng
authors.affiliation: Key Laboratory of Separation Sciences for Analytical Chemistry, National Chromatographic R&A Center, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China.
authors.forename: Hanfa
authors.initials: H
authors.lastname: Zou
mesh_terms: D053148:Caspase 3 / Q000032:analysis / Q000737:chemistry*; D006801:Humans; D019169:Jurkat Cells; D008956:Models, Chemical; D059748:Proteolysis; D013379:Substrate Specificity
chemical_list: C505407:CASP3 protein, human; D053148:Caspase 3