How conformational dynamics shape hydrogen–deuterium exchange isotopic envelopes

Hydrogen–deuterium exchange mass spectrometry (HDX–MS) measures peptide-level mass distributions, whereas exchange is controlled microscopically by fluctuations in the exposure of individual backbone amides. In this work, exchange is represented in terms of integrated conformation-dependent instantaneous exchange rates that are accumulated along stochastic conformational trajectories, and peptide uptake statistics are given by appropriate survival probabilities, which obey Feynman–Kac equations. The framework shows that centroid uptake curves depend only on single-residue survival probabilities, whereas the full envelope can present signatures reflecting correlated exchange histories through pair and higher-order subset survival probabilities. If conformational fluctuations are independent, the resulting envelopes approach a Poisson binomial distribution. When residues share conformational histories, correlations can produce broadened and possibly multimodal envelopes. Whether such features are visible depends on the timescales of relevant conformational changes. The same formalism naturally extends to non-equilibrium and pulse labeling experiments by changing the ensemble of trajectories being averaged. Finally, the effect of heterogeneous back exchange is described as a mapping from the post-labeling exchange statistics to the experimentally measured mass spectrum. The result is a general bridge between microscopic conformational dynamics and experimentally observed HDX–MS envelopes.