Hydrogen-deuterium exchange (HDX) is a technique that measures the exchange of protein hydrogens for deuteriums in a D2O-containing buffer, providing readout of the structural dynamics. Histidine hydrogen- deuterium exchange mass spectrometry (His-HDX-MS) is a variation of this technique that measures the slow HDX of imidazole C2 hydrogens of histidines. This measurement, when accompanied by pH titration, provides both pKas and half-lives (t1/2) of the HDX reaction for individual histidine residues in proteins. The pKa and t1/2 values indicate the electrostatic environment and the degree of side-chain solvent accessibility of the histidine residues, respectively. Herein we describe an experimental protocol to characterize rhodopsin by His-HDX-MS. This technique can be used to monitor different states of rhodopsin and might be useful for monitoring longtime scale events in other GPCRs.
CITATION STYLE
Lodowski, D. T., & Miyagi, M. (2015). Analysis of conformational changes in rhodopsin by histidine hydrogen-deuterium exchange. In Rhodopsin: Methods and Protocols (pp. 123–132). Springer New York. https://doi.org/10.1007/978-1-4939-2330-4_9
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