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A Pan Photoaffinity Probe for Detecting Active Forms of Matrix Metalloproteinases

Authors

  • Catherine Nury,

    1. CEA (Commissariat à l'Energie Atomique), Service d'Ingénierie Moléculaire des Protéines, (SIMOPRO), CEA-Saclay 91191 Gif/Yvette Cedex (France)
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  • Bertrand Czarny,

    1. CEA (Commissariat à l'Energie Atomique), Service d'Ingénierie Moléculaire des Protéines, (SIMOPRO), CEA-Saclay 91191 Gif/Yvette Cedex (France)
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  • Evelyne Cassar-Lajeunesse,

    1. CEA (Commissariat à l'Energie Atomique), Service d'Ingénierie Moléculaire des Protéines, (SIMOPRO), CEA-Saclay 91191 Gif/Yvette Cedex (France)
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  • Dr. Dimitris Georgiadis,

    1. Laboratory of Organic Chemistry, University of Athens, Panepistimiopolis, Zografou, 15771 Athens (Greece)
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  • Dr. Sarah Bregant,

    1. CEA (Commissariat à l'Energie Atomique), Service d'Ingénierie Moléculaire des Protéines, (SIMOPRO), CEA-Saclay 91191 Gif/Yvette Cedex (France)
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  • Dr. Vincent Dive

    Corresponding author
    1. CEA (Commissariat à l'Energie Atomique), Service d'Ingénierie Moléculaire des Protéines, (SIMOPRO), CEA-Saclay 91191 Gif/Yvette Cedex (France)
    • CEA (Commissariat à l'Energie Atomique), Service d'Ingénierie Moléculaire des Protéines, (SIMOPRO), CEA-Saclay 91191 Gif/Yvette Cedex (France)
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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 P1′ 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.

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