SARS-CoV-2主蛋白酶的表达、纯化和一种新型晶体结构的研究  

Expression and Purification of SARS-CoV-2 Main Protease and Study of Novel Crystal Structure

作　　者：刘雪静 赵文聪 高欣欣 陈思旭 张博涛 王君帅 李朝阳[2] 温一帆 高靓 曹鹏 

机构地区：[1]北京工业大学材料科学与工程学院,北京 [2]北京工业大学化学与生命科学学院,北京

出　　处：《生物医学》2024年第2期229-239,共11页Hans Journal of Biomedicine

摘　　要：新型冠状病毒SARS-CoV-2的主蛋白酶(Main Protease, Mpro),也称为3-糜蛋白酶样蛋白酶(3CLpro),在病毒复制的过程中发挥核心作用。Mpro的结构和功能在β-冠状病毒中相对保守,在人体内没有同源蛋白,因此是抗病毒药物开发的关键靶点。本研究完成了SARS-CoV-2 Mpro的原核表达与纯化,获得具有高纯度和高均一性的蛋白质样品。利用气相扩散坐滴法开展了Mpro的结晶筛选,获得单晶体后将其浸泡在由谷胱甘肽包裹的金团簇纳米材料溶液中,成功解析了仅有第156位半胱氨酸结合一个金离子的一个新型X射线衍射晶体结构。结构分析发现,Mpro蛋白的Cys156结合金离子后Asp153~Cys156区域的构象发生了~0.6 Å的偏移,推测该构象变化进一步通过变构效应抑制Mpro的生物功能,为新冠病毒的纳米型抑制剂的研发提供了新的信息。The main protease (Mpro) of the novel coronavirus SARS-CoV-2, also known as 3C-like protease (3CLpro), plays a central role in the virus replication process. The structure and function of Mpro are relatively conserved in β-coronaviruses and there is no homologous protein in human, making it a key target for antiviral drug development. In this study, we successfully completed the prokaryotic expression and purification of SARS-CoV-2 Mpro, obtaining a high-purity and high-homo- geneity protein sample. Crystallization screening of Mpro was carried out using the vapor-diffusion sitting-drop method. After obtaining single crystals, they were soaked in a solution of the nano-materialglutathione-protected Au clusters, resulting in a novel X-ray diffraction crystal structure with only one cysteine at position 156 binding to one Au(I) ion. Structural analysis revealed a ~0.6 Å displacement in the conformation of the Asp153~Cys156 region of the Mpro protein upon binding of the Cys156 to the gold ion, suggesting that this conformational change further inhibits the biological function of Mpro through an allosteric effect, providing new information for the development of novel coronavirus inhibitors.

关 键 词：SARS-CoV-2 M^pro 金团簇 蛋白质表达与纯化 X射线晶体结构 

分 类 号：R37[医药卫生—病原生物学]