Combining Ion Mobility Spectrometry with Hydrogen-Deuterium Exchange and Top-Down MS for Peptide Ion Structure Analysis

The gas-phase conformations of electrosprayed ions of the model peptide KKDDDDIIKIIK have been examined by ion mobility spectrometry (IMS) and hydrogen deuterium exchange (HDX)-tandem mass spectrometry (MS/MS) techniques. [M+4H]⁴⁺ ions exhibit two conformers with collision cross sections of 418 Ų and 471 Ų. [M+3H]³⁺ ions exhibit a predominant conformer with a collision cross section of 340 Ų as well as an unresolved conformer (shoulder) with a collision cross section of ~367 Ų. Maximum HDX levels for the more compact [M+4H]⁴⁺ ions and the compact and partially-folded [M+3H]³⁺ ions are ~12.9, ~15.5, and ~14.9, respectively. Ion structures obtained from molecular dynamics simulations (MDS) suggest that this ordering of HDX level results from increased charge-site/exchange-site density for the more compact ions of lower charge. Additionally, a new model that includes two distance calculations (charge site to carbonyl group and carbonyl group to exchange site) for the computer-generated structures is shown to better correlate to the experimentally determined per-residue deuterium uptake. Future comparisons of IMS-HDX-MS data with structures obtained from MDS are discussed with respect to novel experiments that will reveal the HDX rates of individual residues. Graphical Abstract

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