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Détection et identification de sélénoprotéines par électrophorèse sur gel associée aux spectrométries de masse atomique et moléculaire

Guillaume Ballihaut 1
1 LCABIE - Laboratoire de Chimie Analytique Bio-Inorganique et Environnement
IPREM - Institut des sciences analytiques et de physico-chimie pour l'environnement et les materiaux
Abstract : Selenium is a trace element known for its necessity in organisms' development and its beneficial roles in human health. Research in the past five years strengthened the idea that these properties are largely due to the synthesis of selenoproteins in living organisms. Classical methods in proteomics are not adapted to identify minor selenoproteins. In this PhD work, more sensitive and more specific methods complementary to proteomics have been developed for their detection and identification. A stable selenized protein standard has been produced to develop these methods. The development of laser ablation - Inductively Coupled Plasma - Mass spectrometry coupling (LA-ICP-MS) lead to the detection of selenoproteins after separation by polyacrylamide gel electrophoresis (PAGE). Conventional device of laser ablation has been used and then improved with a new device using femtoseconde laser with ultrafast scanning for more sensitive detection of selenoproteins in biological samples. Once detected, selenoproteins have been identified by molecular mass spectrometry. For this purpose selenopeptids from enzymatic digestion of selenoproteins have been detected first by nanochromatography coupled to ICP-MS and sequenced then by means of by nanochromatography coupled to electrospray ionisation tandem mass spectrometry (nanoHPLC-ESI-MS/MS). LA-ICP-MS device has been successful for detection of new selenoproteins in Desulfococcus multivorans bacteria. This identification procedure has been validated on two purified selenoproteins thioredoxin reductase and carboxymathylated glutathion peroxidase. This developed methodology will lead to identification of selenoproteins and a better understanding of the physiological role of these molecules in numerous living organisms.
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Submitted on : Tuesday, January 3, 2012 - 7:10:52 PM
Last modification on : Friday, January 15, 2021 - 9:23:17 AM
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  • HAL Id : tel-00656273, version 1



Guillaume Ballihaut. Détection et identification de sélénoprotéines par électrophorèse sur gel associée aux spectrométries de masse atomique et moléculaire. Chimie analytique. Université de Pau et des Pays de l'Adour, 2007. Français. ⟨tel-00656273⟩



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