Anal Chem. 2020 May 5;92(9):6637-6644. doi: 10.1021/acs.analchem.0c00463. Epub 2020 Apr 14.
A new covalent labeling (CL) reagent based on an α,β-unsaturated carbonyl scaffold has been developed for studying protein structure and protein-protein interactions when coupled with mass spectrometry. We show that this new reagent scaffold can react with up to 13 different types of residues on protein surfaces, thereby providing excellent structural resolution. To illustrate the value of this reagent scaffold, it is used to identify the residues involved in the protein-protein interface that is formed upon Zn(II) binding to the protein β-2-microglobulin. The modular design of the α,β-unsaturated carbonyl scaffold allows facile variation of the functional groups, enabling labeling kinetics and selectivity to be tuned. Moreover, by introducing isotopically enriched functional groups into the reagent structure, labeling sites can be more easily identified by MS and MS/MS. Overall, this reagent scaffold should be a valuable CL reagent for protein higher order structure characterization by MS.
一种基于α,β-不饱和羰基支架的新共价标记 (CL) 试剂已被开发出来，用于与质谱结合研究蛋白质结构和蛋白质-蛋白质相互作用。我们表明，这种新的试剂支架可以与蛋白质表面上多达 13 种不同类型的残基反应，从而提供出色的结构分辨率。为了说明这种试剂支架的价值，它被用于鉴定锌 (II) 与蛋白质β-2-微球蛋白结合时形成的蛋白质-蛋白质界面涉及的残基。α,β-不饱和羰基支架的模块化设计允许轻松改变官能团，从而可以调整标记动力学和选择性。此外，通过在试剂结构中引入同位素富集的官能团，可以更容易地通过 MS 和 MS/MS 鉴定标记位点。总的来说，这种试剂支架应该是通过 MS 对蛋白质高级结构进行特征描述的有价值的 CL 试剂。
