Structural biology has recently been revolutionized by cryo-electron microscopy, which has the capability of revealing structures are large molecular complexes at high resolution. Combined with native mass spectrometry to understand stoichiometries, and often protein cross-linking, structures of several large protein machines have been elucidated. It is our view that these static structures are just the starting point to understand the dynamic functions of large protein complexes. HDXMS is an ideal approach to combine with cryo-EM to reveal dynamic information. We have been testing the limits of the Synapt G2Si to analyze proteins and protein complexes approaching 200kD. The critical role of the ion mobility capabilities of the Synapt G2Si for resolving peptide mass envelope overlaps will be demonstrated. We have also directly compared extended timescale (msec) molecular dynamics simulations to HDXMS experimental results to understand the timescale of motions that are sampled by HDXMS experiments in the fast limit. These comparisons reveal that exchange times under 2 min largely sample amides that are not stably hydrogen-bonded, but are often involved in functional dynamics such as allosteric transitions. The results suggest that for complexes too large for NMR, HDXMS is an ideal experiment for revealing msec motional changes involved in functionally important dynamic allostery.

Hydrogen-deuterium exchange in large complexes: How big can we go and what can we learn?

2019 Fall National Meeting

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