Enzyme-Coupled Nanoparticles-Assisted Laser Desorption Ionization Mass Spectrometry for Searching for Low-Mass Inhibitors of Enzymes in Complex Mixtures |
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Authors: | Aleksander Salwiński David Da Silva Raphaël Delépée Benoît Maunit |
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Institution: | 1. University of Orléans, CNRS, ICOA, UMR 7311, 45067, Orleans, France 2. Normandy University, UCBN, UR ABTE EA 4651, 14032, Caen, France
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Abstract: | In this report, enzyme-coupled magnetic nanoparticles (EMPs) were shown to be an effective affinity-based tool for finding specific interactions between enzymatic targets and the low-mass molecules in complex mixtures using classic MALDI-TOF apparatus. EMPs used in this work act as nonorganic matrix enabling ionization of small molecules without any interference in the low-mass range (enzyme-coupled nanoparticles-assisted laser desorption ionization MS, ENALDI MS) and simultaneously carry the superficial specific binding sites to capture inhibitors present in a studied mixture. We evaluated ENALDI approach in two complementary variations: ‘ion fading’ (IF-ENALDI), based on superficial adsorption of inhibitors and ‘ion hunting’ (IH-ENALDI), based on selective pre-concentration of inhibitors. IF-ENALDI was applied for two sets of enzyme–inhibitor pairs: tyrosinase–glabridin and trypsin–leupeptin and for the real plant sample: Sparrmannia discolor leaf and stem methanol extract. The efficacy of IH-ENALDI was shown for the pair of trypsin–leupeptin. Both ENALDI approaches pose an alternative for bioassay-guided fractionation, the common method for finding inhibitors in the complex mixtures. Figure ? |
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