title: Development of a novel assay for proprotein converting enzyme activity on a multiplex bead-based array system.
abstract: We describe the development of a novel, robust assay system for determining the changes in activity of proprotein converting enzymes. An assay for prolyl oligopeptidase (POP) activity was constructed using a peptide-streptavidin substrate coupled to magnetic microspheres and cleavage was detected by loss of streptavidin on the MAGPIX reader. Test analysis of postmortem pituitary extracts from schizophrenia patients showed an increase in POP activity compared to controls. The results were validated using both fluorometric and Western blot analyses for POP activity and immunoreactivity, respectively. The assays can be multiplexed for measuring the activity of multiple proprotein cleaving enzymes simultaneously in laboratory and clinical settings and should add valuable new information for conditions such as neuropsychiatric diseases, diabetes, endocrine dysfunction, and cancer, where effects on proteolysis of biologically active peptides play a key role.
authors: Department of Chemical Engineering and Biotechnology, University of Cambridge, Cambridge, UK.
authors: Andreas
authors: A
authors: Albrecht
authors: Hassan
authors: H
authors: Rahmoune
authors: Kristi
authors: K
authors: Leedjärv
authors: Thomas
authors: T
authors: Knorpp
authors: Thomas
authors: T
authors: Joos
authors: Pawel
authors: P
authors: Stocki
authors: Paul C
authors: PC
authors: Guest
authors: Sabine
authors: S
authors: Bahn
mesh_terms: D015153:Blotting, Western; D016022:Case-Control Studies; D003710:Demography; D057075:Enzyme Assays / Q000379:methods*; D005260:Female; D005453:Fluorescence; D006801:Humans; D008297:Male; D008863:Microspheres; D008875:Middle Aged; D010902:Pituitary Gland / Q000201:enzymology; D000085822:Prolyl Oligopeptidases; D040081:Protein Array Analysis / Q000379:methods*; D012559:Schizophrenia / Q000201:enzymology; D012697:Serine Endopeptidases / Q000378:metabolism*
chemical_list: D012697:Serine Endopeptidases; C000705287:PREPL protein, human; D000085822:Prolyl Oligopeptidases
keywords: Biomedicine; Microarrays; Multiplex protein assay; Proprotein converting enzyme; Schizophrenia; Technology