title: Analysis of conformational changes in rhodopsin by histidine hydrogen-deuterium exchange.
abstract: 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 pK as and half-lives (t 1/2) of the HDX reaction for individual histidine residues in proteins. The pK a and t 1/2 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.
authors.affiliation: Case Center for Proteomics and Bioinformatics, Case Western Reserve University, 10900 Euclid Ave., Cleveland, OH, 44106-4965, USA.
authors.forename: David T
authors.initials: DT
authors.lastname: Lodowski
authors.forename: Masaru
authors.initials: M
authors.lastname: Miyagi
mesh_terms: D003903:Deuterium / Q000737:chemistry*; D041961:Deuterium Exchange Measurement / Q000379:methods*; D006639:Histidine / Q000737:chemistry*; D006859:Hydrogen / Q000737:chemistry*; D013058:Mass Spectrometry; D011487:Protein Conformation
chemical_list: D006639:Histidine; D006859:Hydrogen; D003903:Deuterium