Advances in Mass Spectrometry‐Based Structural Proteomics: Development of HDX‐MS and XL‐MS Techniques from Recombinant Protein to Cellular Systems

Abstract

Mass spectrometry (MS) is a central analytical technique used to study proteins and biomolecules. It measures mass‐to‐charge ratio of ions to identify and quantify molecules in simple and complex mixtures. Technological advancement in instrumentation, sample preparation methodologies, and data analysis workflows continue to push the capabilities of MS to answer more complicated questions and vice versa. Structural proteomics uses MS‐based methodologies to characterize protein structure. Specifically, but not limited to, hydrogen deuterium exchange MS (HDX‐MS) and crosslinking MS (XL‐MS) are complementary techniques that capture the structural plasticity inherent to proteins in solution. This review is intended to present recent progress in HDX‐MS and XL‐MS that have allowed these techniques to be used not only for simple recombinant protein systems but with complex cellular systems.

Significance of Histidine Hydrogen–Deuterium Exchange Mass Spectrometry in Protein Structural Biology

“…Additionally, a highly efficient histidine-containing peptide enrichment method is available [34], which significantly simplifies proteome samples and enhances the identification of His-containing peptides. For these reasons, His-HDX-MS should be applied more to proteome-scale samples in the near future, as also noted by Courouble and colleagues in their recent review paper [44]. Lastly, there is currently no computational tool available for automating the data analysis step.…”

“…The most commonly used mass spectrometry technique when studying protein structure and dynamics is amide-HDX-MS [43][44][45], which monitors the HDX rates of the backbone amide. The major technical challenge of amide-HDX-MS is the fast back exchange from deuterium to proton, which can occur during protein digestion and mass spectrometry analysis.…”