title: Identification of prolyl oligopeptidase as a cyclosporine-sensitive protease by screening of mouse liver extracts.
abstract: Cyclosporine A (CsA) is a cyclic undecapeptide well known for its ability to prevent rejection episodes after organ transplantation via gain-of-function. Therefore, biomedical studies on CsA have been focused on both immunosuppressive properties and binding to the biocatalytically-active immune receptors, the cyclophilins. Much less attention has been spent on effects of cyclosporines on the biological function of other proteins. We used a 9-mer fluorescence-quenched peptide library with defined sequences to identify cyclosporine-sensitive proteolysis in mouse liver extracts. A highly soluble [d-Ser]8-CsA derivative was utilized to avoid drug precipitation at extended incubation times. Analysis of the time courses of proteolysis revealed 15 out of 360 peptide sequences where proteolysis exhibited marked sensitivity to the cyclosporine derivative. As a first example, a collagen-derived substrate was selected from those hits to identify the targeted proteolytic pathway. After purification from mouse liver extracts, prolyl oligopeptidase (EC 3.4.21.26) could be identified as a protease sensitive to submicromolar concentrations of cyclosporines. Surprisingly, in a series of cyclosporine derivatives an inverse relationship was found between the inhibition of prolyl oligopeptidase and inhibition of cyclophilin A.
authors.affiliation: Max Planck Research Unit for Enzymology of Protein Folding, Weinbergweg 22, D-06120 Halle, Germany.
authors.forename: Alexander
authors.initials: A
authors.lastname: Rentzsch
authors.forename: Viktoria
authors.initials: V
authors.lastname: Kahlert
authors.forename: Günther
authors.initials: G
authors.lastname: Jahreis
authors.forename: Bernhard
authors.initials: B
authors.lastname: Schlott
authors.forename: Mike
authors.initials: M
authors.lastname: Schutkowski
authors.forename: Miroslav
authors.initials: M
authors.lastname: Malešević
authors.forename: Gunter
authors.initials: G
authors.lastname: Fischer
mesh_terms: D000818:Animals; D021983:Cyclophilins / Q000378:metabolism; D016572:Cyclosporine / Q000378:metabolism*; D006801:Humans; D007166:Immunosuppressive Agents / Q000378:metabolism*; D008110:Liver Extracts / Q000737:chemistry / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D019151:Peptide Library; D000085822:Prolyl Oligopeptidases; D059748:Proteolysis; D012697:Serine Endopeptidases / Q000302:isolation & purification* / Q000378:metabolism*
chemical_list: D007166:Immunosuppressive Agents; D008110:Liver Extracts; D019151:Peptide Library; D016572:Cyclosporine; D012697:Serine Endopeptidases; C000705287:PREPL protein, human; D000085822:Prolyl Oligopeptidases; D021983:Cyclophilins