Department of Chemistry, York University, Toronto M3J 1P3, Canada.
Department of Chemistry, York University, Toronto M3J 1P3, Canada; Centre for Research of Biomolecular Interactions, York University, Toronto M3J 1P3, Canada; Centre for Research in Mass Spectrometry, York University, Toronto M3J 1P3, Canada.
Methods. 2018 Jul 15;144:27-42. doi: 10.1016/j.ymeth.2018.04.023. Epub 2018 Apr 26.
Hydrogen/deuterium exchange (HDX) mass spectrometry (MS) emerged as a tool for biochemistry and structural biology around 25 years ago. It has since become a key approach for studying protein dynamics, protein-ligand interactions, membrane proteins and intrinsically disordered proteins (IDPs). In HDX labeling, proteins are exposed to deuterated solvent (usually DO) for a variable 'labeling time', resulting in isotope exchange of unprotected labile protons on the amide backbone and amino acid side chains. By comparing the levels of deuterium uptake in different regions of a protein, information on conformational and dynamic changes in the system can be acquired. When coupled with MS, HDX is suitable for probing allosteric effects in catalysis and ligand binding, epitope mapping, validation of biosimilars, drug candidate screening and mapping membrane-protein interactions among many other bioanalytical applications. This review introduces HDX-MS via a brief description of HDX-MS development, followed by an overview of HDX theory and ultimately an outline of methods and procedures involved in performing HDX-MS experiments.
氢/氘交换（HDX）质谱（MS）大约在 25 年前作为一种生物化学和结构生物学工具出现。从那时起，它已成为研究蛋白质动力学、蛋白质-配体相互作用、膜蛋白和天然无序蛋白（IDP）的关键方法。在 HDX 标记中，将蛋白质暴露于氘化溶剂（通常为 DO）中进行可变的“标记时间”，导致酰胺骨架和氨基酸侧链上未保护的不稳定质子的同位素交换。通过比较蛋白质不同区域的氘吸收水平，可以获得系统中构象和动态变化的信息。当与 MS 结合使用时，HDX 适用于探测催化和配体结合中的变构效应、表位作图、生物类似物的验证、候选药物筛选以及映射膜蛋白相互作用等许多其他生物分析应用。本综述通过简要描述 HDX-MS 的发展，介绍了 HDX-MS，随后概述了 HDX 理论，最终概述了进行 HDX-MS 实验所涉及的方法和程序。
