Supported inhibitor for fishing lipases in complex biological media and mass spectrometry identification

Vincent Delorme, Brigitt Raux, Rémy Puppo, Julien Leclaire, Jean-François Cavaliera, Sylvain Marc, Pavan-Kumar Kamarajugadd, Gérard Buono, Frédéric Fotiadu, Stéphane Canaan, Frédéric Carrière
Biochimie. 2014 Dec; 107: 124-134
DOI: 10.1016/j.biochi.2014.07.015

A synthetic phosphonate inhibitor designed for lipase inhibition but displaying a broader range of activity was covalently immobilized on a solid support to generate a function-directed tool targeting serine hydrolases. To achieve this goal, straightforward and reliable analytical techniques were developed, allowing the monitoring of the solid support’s chemical functionalization, enzyme capture processes and physisorption artifacts. This grafted inhibitor was tested on pure lipases and serine proteases from various origins, and assayed for the selective capture of lipases from several complex biological extracts. The direct identification of captured enzymes by mass spectrometry brought the proof of concept on the efficiency of this supported covalent inhibitor. The features and limitations of this « enzyme-fishing » proteomic tool provide new insight on solid-liquid inhibition process.

Auteurs Bordeaux Neurocampus