Structure Characterization of a Disordered Peptide Using In-Droplet Hydrogen/Deuterium Exchange Mass Spectrometry and Molecular Dynamics

In-droplet hydrogen/deuterium exchange (HDX)-mass spectrometry (MS) experiments have been conducted for peptides of highly varied conformational type. A new model is presented that combines the use of protection factors (PF) from molecular dynamics (MD) simulations with intrinsic HDX rates (<i>k</i> _int) to obtain a structure-to-reactivity calibration curve. Using the model, the relationship of peptide structural flexibility and HDX reactivity for different peptides is elucidated. Additionally, the model is used to describe the degree of conformational flexibility and structural bias for the disease-relevant Nt17 peptide; although highly flexible, intrinsically primed for facile conversion to α-helical conformation upon binding with molecular partners imparts significant in-droplet HDX protection for this peptide. In the future, a scale may be developed whereby HDX reactivity is predictive of the degree of structural flexibility and bias (propensity to form 2° structural elements such as α-helix, β-sheet, and β-turn) for intrinsically disordered regions (IDRs). Such structural resolution may ultimately be used for high-throughput screening of IDR structural transformation(s) upon binding of ligands such as drug candidates.