Enhancement of the thermostability of mouse claudin-3 on complex formation with the carboxyl-terminal region of Clostridium perfringens enterotoxin improves crystal quality
Enhancement of the thermostability of mouse claudin-3 on complex formation with the carboxyl-terminal region of Clostridium perfringens enterotoxin improves crystal quality
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增强小鼠claudin-3与产气荚膜梭菌肠毒素羧基末端区域形成复合物的热稳定性，提高晶体质量
DOI：
10.1107/s2053230x18002005
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发表时间：
2018
期刊：
影响因子：
--
通讯作者：
Irie Katsumasa
中科院分区：
文献类型：
--
作者：
Nakamura Shun;Fujiyoshi Yoshinori;Irie Katsumasa
紧密连接作为一种物理屏障或细胞旁途径调节物质通过细胞间隙的渗透，并在维持内环境中起重要作用。紧密连接蛋白是一种四跨膜蛋白，是紧密连接的关键组成部分。在哺乳动物中，已经鉴定了27种claudin亚型，每种亚型都与特定的亚型相互作用。虽然已经确定了几种亚型的晶体结构，但亚型特异性的分子机制仍不清楚。在这里，小鼠密蛋白-3（mCldn 3）与产气荚膜梭菌肠毒素（C-CPE）的C-末端区域复合结晶，用于其他密蛋白亚型的结构分析。单独使用mCldn 3难以结晶，但与C-CPE形成复合物增强了mCldn 3的热稳定性并促进了其结晶。将S313 A突变引入C-CPE进一步改善了其热稳定性，衍射数据集的分辨率极限从野生型复合物的8 μ m提高到S313 A突变体复合物的4.7 μ m。
Tight junctions regulate substance permeation through intercellular spaces as a physical barrier or a paracellular pathway, and play an important role in maintaining the internal environment. Claudins, which are tetraspan-transmembrane proteins, are pivotal components of tight junctions. In mammals 27 claudin subtypes have been identified, each of which interacts with specific subtypes. Although the crystal structures of several subtypes have been determined, the molecular mechanisms underlying subtype specificity remain unclear. Here, mouse claudin-3 (mCldn3) was crystallized in complex with the C-terminal region of Clostridium perfringens enterotoxin (C-CPE) for the structural analysis of an additional claudin subtype. mCldn3 alone was difficult to crystallize, but complex formation with C-CPE enhanced the thermostability of mCldn3 and facilitated its crystallization. The introduction of an S313A mutation into C-CPE further improved its thermostability, and the resolution limits of the diffraction data sets improved from 8 Å for the wild-type complex to 4.7 Å for the S313A mutant complex.