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Hydrogen/Deuterium Exchange Mass Spectrometry for the Structural Analysis of Detergent-Solubilized Membrane Proteins

Abstract : Integral membrane proteins are involved in numerous biological functions and represent important drug targets. Despite their abundance in the human proteome, the number of integral membrane protein structures is largely underrepresented in the Protein Data Bank. The challenges associated with the biophysical characterization of such biological systems are well known. Most structural approaches, including X-ray crystallography, SAXS, or mass spectrometry (MS), require the complete solubilization of membrane proteins in aqueous solutions. Detergents are frequently used for this task, but may interfere with the analysis, as is the case with MS. The use of "MS-friendly" detergents, such as non-ionic alkyl glycoside detergents, has greatly facilitated the analysis of detergent-solubilized membrane proteins. Here, we describe a protocol, which we have successfully implemented in our laboratory to study the structure and dynamics of detergent-solubilized integral membrane proteins by Hydrogen/Deuterium eXchange and Mass Spectrometry (HDX-MS). The procedure does not require detergent removal prior to MS analysis, instead taking advantage of the ultra-high pressure chromatographic system to separate deuterated peptides from "MS-friendly" detergents.
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Submitted on : Friday, July 31, 2020 - 3:59:38 PM
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Darragh O’brien, Véronique Hourdel, Alexandre Chenal, Sébastien Brier. Hydrogen/Deuterium Exchange Mass Spectrometry for the Structural Analysis of Detergent-Solubilized Membrane Proteins. Springer. Expression, Purification, and Structural Biology of Membrane Proteins, 2127, Humana, New York, NY, pp.339-358, 2020, Methods in Molecular Biology, 978-1-0716-0372-7. ⟨10.1007/978-1-0716-0373-4_22⟩. ⟨pasteur-02559306⟩

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