title: Profiling treatment-specific post-translational modifications in a complex proteome with subtractive substrate phage display.
abstract: Proteolytic activation of zymogens or controlled degradation of inhibitory factors is part of a major regulatory system on the post-translational level to regulate treatment induced cellular stress responses. The identification of differential activity based substrates is thus of high interest to prioritize and validate candidate targets for drug discovery. Here we present a novel subtractive substrate phage display screening method for the selection of treatment induced post-translational peptide modifications in complex proteomes. We investigated this approach with tumor cells in response to a protease activating anticancer treatment modality using subtractive and iterative screening of cellular extracts derived from control and treated cells. Specific phage were identified that served as substrates for proteolytic activities in response to treatment related activity changes and could be distinguished from substrates for unspecific proteolytic background activities. Novel, selected peptide substrates were investigated in vitro and in vivo and showed high substrate specificity and functional biological significance.
authors.affiliation: Department of Radiation Oncology, University Hospital Zurich, Zurich, Switzerland.
authors.forename: Angela
authors.initials: A
authors.lastname: Tenzer
authors.forename: Barbara
authors.initials: B
authors.lastname: Hofstetter
authors.forename: Christelle
authors.initials: C
authors.lastname: Sauser
authors.forename: Stephan
authors.initials: S
authors.lastname: Bodis
authors.forename: August P
authors.initials: AP
authors.lastname: Schubiger
authors.forename: Christophe
authors.initials: C
authors.lastname: Bonny
authors.forename: Martin
authors.initials: M
authors.lastname: Pruschy
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D020169:Caspases / Q000378:metabolism; D002478:Cells, Cultured; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D006801:Humans; D051379:Mice; D019151:Peptide Library; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D020543:Proteome / Q000032:analysis*; D015203:Reproducibility of Results
chemical_list: D019151:Peptide Library; D010455:Peptides; D020543:Proteome; D020169:Caspases