title: Identification of Protease Cleavage Sites by Charge-Based Enrichment of Protein N-Termini.
abstract: Differential proteolytic processing, for example by matrix metalloproteases (MMPs), has been recognized as an important hallmark in numerous pathological conditions. One crucial challenge in the present studies of proteases is system-wide identification of endogenous biological substrates. In this chapter, we highlight a robust method for the identification of bioactive substrates and their sites of MMP cleavage, as well as by other proteases and peptidases, in a system-wide manner. This approach enriches for putative protein N-termini by removal of internal peptides using a charge reversal strategy. In addition, this straightforward method can be used in combination with gel-based pre-separation of proteins to allow better estimation of the molecular weight of the identified cleavage product of a given bioactive substrate.
authors.affiliation: Institute of Molecular Medicine and Cell Research, University of Freiburg, 79104, Freiburg, Germany.|Department of Genetics and Complex Diseases, Harvard T. H. Chan School of Public Health, Boston, MA, 02115, USA.
authors.forename: Zon W
authors.initials: ZW
authors.lastname: Lai
authors.affiliation: Institute of Molecular Medicine and Cell Research, University of Freiburg, 79104, Freiburg, Germany. oliver.schilling@mol-med.uni-freiburg.de.|BIOSS Centre of Biological Signaling Studies, University of Freiburg, 79104, Freiburg, Germany. oliver.schilling@mol-med.uni-freiburg.de.|German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ), Heidelberg, Germany. oliver.schilling@mol-med.uni-freiburg.de.
authors.forename: Oliver
authors.initials: O
authors.lastname: Schilling
mesh_terms: D020782:Matrix Metalloproteinases / Q000378:metabolism*; D008970:Molecular Weight; D010455:Peptides / Q000737:chemistry; D059748:Proteolysis; D020543:Proteome / Q000737:chemistry*; D040901:Proteomics / Q000379:methods*; D013379:Substrate Specificity; D053719:Tandem Mass Spectrometry
chemical_list: D010455:Peptides; D020543:Proteome; D020782:Matrix Metalloproteinases
keywords: Cleavage; Degradome; Protease; Proteomics; Substrate