title: A pan photoaffinity probe for detecting active forms of matrix metalloproteinases.
abstract: A photoaffinity probe based on the scaffold of a potent broad-spectrum phosphinic peptide inhibitor of matrix metalloproteinases (MMPs) has been developed. A photolabile diazirine group for covalent modification of MMP active forms was incorporated at the P(1) ' position, and a tritium radioactive label for the sensitive detection of MMP covalent adducts by radioimaging was attached. The probe was characterized on seven catalytic domains of human MMPs (MMP-2, -3, -8, -9, -12, -13 and -14) and was found to display nanomolar affinities towards this set of MMPs, covalently modifying them with crosslinking yields varying from 12 to 58 %, thus leading to highly sensitive detection of these MMPs. In a complex proteome complemented with four recombinant MMPs (MMP-2, -9, -12 and -13), this probe enabled their simultaneous detection with a threshold of few femtomoles and low background labelling. Those properties should make this new pan-activity-based MMP probe a valuable tool for the detection of MMP active forms from biological fluids or tissue extracts.
authors.affiliation: CEA (Commissariat à l'Energie Atomique), Service d'Ingénierie Moléculaire des Protéines, (SIMOPRO), CEA-Saclay 91191 Gif/Yvette Cedex, France.
authors.forename: Catherine
authors.initials: C
authors.lastname: Nury
authors.forename: Bertrand
authors.initials: B
authors.lastname: Czarny
authors.forename: Evelyne
authors.initials: E
authors.lastname: Cassar-Lajeunesse
authors.forename: Dimitris
authors.initials: D
authors.lastname: Georgiadis
authors.forename: Sarah
authors.initials: S
authors.lastname: Bregant
authors.forename: Vincent
authors.initials: V
authors.lastname: Dive
mesh_terms: D001386:Azides / Q000737:chemistry; D001389:Azirines / Q000737:chemistry; D020134:Catalytic Domain; D003432:Cross-Linking Reagents / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D008027:Light; D020782:Matrix Metalloproteinases / Q000737:chemistry / Q000378:metabolism*; D015336:Molecular Probe Techniques; D019920:Photoaffinity Labels / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D001386:Azides; D001389:Azirines; D003432:Cross-Linking Reagents; D019920:Photoaffinity Labels; D020782:Matrix Metalloproteinases