Mass spectrometric determination of disulfide bonds and free cysteine in grass carp IgM isoforms
Mass spectrometric determination of disulfide bonds and free cysteine in grass carp IgM isoforms
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质谱法测定草鱼IgM亚型中二硫键和游离半胱氨酸
DOI：
10.1016/j.fsi.2019.10.051
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发表时间：
2019
影响因子：
4.7
通讯作者：
Geng Hui
中科院分区：
文献类型：
--
作者：
Su Yiling;Wang Bing;Zhang Ying;Ruan Zilun;Bai Hao;Wan Jian;Xu Chen;Li Guoqi;Wang Shengqiang;Ai Hui;Xiong Li;Geng Hui
二硫键是构建Ig结构和维持Ig生物学功能的基础。本文采用液相色谱-电喷雾串联质谱（LC-ESI-MS/MS）分析了草鱼IgM的三种异构体（单体、二聚体/三聚体和四聚体）中的二硫键和游离半胱氨酸。结果表明，cys574在单体IgM中与聚合IgM中c末端末端的残基状态存在显著差异，cys574在单体IgM中为游离巯基，而在二聚体/三聚体和四聚体IgM中形成二硫键。在单体、二聚体/三聚体和四聚体IgM中，CH1~CH4和CL1结构域的5个链内二硫键以及1个H-H和1个H-L链间二硫键也被观察到并显示出相同的连通性。这些发现代表了草鱼IgM二硫键的首次实验分配，并揭示了草鱼IgM异构体的形成是由于连接c末端末端cys574残基的替代二硫键。
Disulfide bonds are fundamental in establishing Ig structure and maintaining Ig biological function. Here, we analysed disulfide bonds and free cysteine in three grass carp IgM isoforms (monomeric, dimeric/trimeric, and tetrameric IgM) by liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS). The results revealed that Cys574residue status at the C-terminal tail differed substantially in monomeric IgM in comparison with polymeric IgM, Cys574was found as free thiol in monomeric IgM, while it formed disulfide linkages in dimeric/trimeric and tetrameric IgM. Five intra-chain disulfide bonds in the CH1~CH4 and CL1 domains, as well as one H–H and one H-L inter-chain disulfide linkages, were also observed and shown identical connectivity in monomeric, dimeric/trimeric, and tetrameric IgM. These findings represent the first experimental assignments of disulfide linkages of grass carp IgM and reveal that grass carp IgM isoform formation is due to alternative disulfide bonds connecting the Cys574residue at the C-terminal tail.