Multi-Ensemble Reweighting Enables Structural Interpretation of Differential HDX-MS

Multi-ensemble reweighting enables quantitative structural interpretation of differential HDX-MS despite >50% deuterium loss, capturing alternating-access transitions in LeuT and NCX_Mj.

• Why it matters: Differential HDX-MS is widely used, but deuterium loss during preparation and analysis often exceeds 50%, limiting existing reweighting methods and making experimental loss correction impractical.
• What they did: The method models shared, peptide-dependent deuterium loss without requiring its characterization, validated on synthetic benchmarks and applied to wild-type and Y268A LeuT, then to substrate-induced occlusion in NCX_Mj.
• The result: It improves agreement with X-ray structures, outperforms conventional reweighting with loss-corrected data, recovers experimentally supported structural changes, and ranks AlphaFold3-derived ensembles comparably to simulation-derived reweighting.