The relatively short time scales employed during gas-phase HDX allow for selective labeling of fast exchanging (i.e. side chain) hydrogens. In the presence of secondary structures, such as alpha-helices, amide hydrogens may be protected from exchange. In the case of the SH3 domain, gas-phase HDX detected two discretely folded states. Binding of peptide PD1R either inhibited necessary motions required to obtain the alternative state or bound exclusively to the predominant state. SH3 and DHFR results highlight the utility of gas-phase HDX to interrogate global changes upon ligand binding that correlate with known changes in protein structural dynamics. Current work focuses on obtaining more complete ECD fragmentation of larger proteins to understand local label variations.
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