HDX-guided EPR spectroscopy to interrogate membrane protein dynamics
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Updated: Sep 3, 2025
Time-resolved ElectroSpray Ionization Hydrogen-deuterium Exchange Mass Spectrometry for Studying Protein Structure and Dynamics
Published on: April 17, 2017
Benjamin J Lane1, Bolin Wang1, Yue Ma1
1Astbury Centre for Structural Molecular Biology, University of Leeds, Leeds LS2 9JT, UK; School of Biomedical Sciences, Faculty of Biological Sciences, University of Leeds, Leeds LS2 9JT, UK.
Abstract:
Solvent accessibilities of and distances between protein residues measured by pulsed-EPR approaches provide high-resolution information on dynamic protein motions. We describe protocols for the purification and site-directed spin labeling of integral membrane proteins. In our protocol, peptide-level HDX-MS is used as a precursor to guide single-residue resolution ESEEM accessibility measurements and spin labeling strategies for EPR applications. Exploiting the pentameric MscL channel as a model, we discuss the use of cwEPR, DEER/PELDOR, and ESEEM spectroscopies to interrogate membrane protein dynamics. For complete details on the use and execution of this protocol, please refer to Wang et al. (2022).
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