Abstract
Hydrogen/deuterium exchange (HDX) is used in protein biophysics to probe folding dynamics, intermolecular interactions, epitope and other mapping. A typical procedure often involves HDX in buffered D2O solution followed by pepsin digestion, and liquid chromatography/electrospray ionization mass spectrometry analysis. In this work, HDX of protein ions was conducted in the ESI source. Both native electrospray droplets of ubiquitin and denatured myoglobin were exposed to D2O vapor in the source region of a Bruker SolariX 12T FTICR-mass spectrometer. Electron capture dissociation was used to assess deuterium incorporation at the residue level. This in-source HDX, on the millisecond-time scale, exchanges side-chain hydrogens and fast-exchanging amides compared to conventional minutes-to-hours HDX of backbone hydrogens in solution with less sample preparation (i.e., no D2O/protein mixing and incubation, no quenching, protein digestion, or LC separation).
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