H5N1禽流感病毒血凝素蛋白的受体结合特异性及其结构基础研究

H5N1 highly pathogenic avian influenza virus has been an important pathogen hazard of poultry industry in China, and its interspecies transmission to human has posed a serious threat to public health. Hemagglutinin (HA) -mediated receptor binding is a major determinant of influenza virus interspecies transmission. In 2013, we determined the complex structures of a mutant HA from a ferret-transmissible H5N1 virus bound to either avian or human receptor sialic acid analogs, providing a structural basis to understand how Q226L substitution alters the receptor binding preference of H5N1 HA. However, there are still many amino acid substitutions which can alter the receptor binding specificity of H5N1 HA, and the molecular mechanisms of these changing have not been elucidated. . Here, we have expressed the HA of an H5N1 virus isolated from Qinghai Lake (QH05-H5) and solved its crystal structure. Our project is to introduce amino acid substitutions into the QH05-H5 to general mutants which may obtain human receptor binding specificity, and screen positive mutants by examining the receptor binding properties of these mutant H5 proteins as well as H5N1 pseudoviruses. We will also solve the complex structures of positive mutants bound to human receptor analogs to provide the structural basises for the H5 adaptations to acquire human receptor binding properties.. Our research will elucidate the molecular mechanisms of interspecies transmission of H5N1 influenza virus, and provide an important theoretical basis for the monitoring and early warning of the pandemic of H5N1 influenza virus in human.

H5N1高致病性禽流感病毒一直是危害我国养禽业的重要病原体，而其跨种间传播所引起的人类感染给公共卫生带来严重威胁。血凝素蛋白（HA）介导的受体结合是流感病毒跨种间传播的主要决定因素。2013年，本实验室解析了可在雪貂中传播的H5N1病毒突变体的HA与受体类似物的复合物结构，阐明了Q226L突变使H5N1 HA获得人源受体结合能力的分子机制。然而，还有很多氨基酸突变能改变H5N1 HA的受体结合特异性，其分子机制尚未阐明。本项目计划在已完成了蛋白表达和结构解析等前期工作的H5N1青海湖分离株HA的基础上，引入各种可能使H5N1 HA获得人源受体结合能力的突变，从蛋白和病毒层面研究这些突变体的受体结合特异性，并利用X-射线晶体学的方法解释其倾向于结合人源受体的结构基础，阐明H5N1流感病毒跨种间传播的分子机制，为监测和预警H5N1流感病毒在人际间的流行提供重要的理论依据。

近年来，H5N1、H5N2、H5N6和H5N8禽流感病毒不时在家禽中引起大规模的禽流感疫情，而其中的H5N1和H5N6禽流感病毒感染人的案例也时有发生，这些H5亚型的禽流感病毒发生跨种间传播感染人类的风险引起了极大的关注。血凝素蛋白（HA）介导的受体结合是流感病毒跨种间传播的主要决定因素。研究表明Q226L突变能使H5N1 HA获得人源受体结合能力，但该突变在天然分离的H5亚型流感病毒中并不存在。因此，其他氨基酸突变对H5亚型HA的受体结合特异性的影响同样值得关注。. 本项目研究了一株天然分离的在病毒水平上能结合人源受体的H5N2禽流感病毒的HA蛋白的受体结合特异性，发现其HA蛋白确实获得了人源受体结合能力，但同时仍保留了对禽源受体的高亲和力。我们分析了该病毒发生人际传播的可能性，并解析了其HA蛋白与禽源/人源受体的复合物晶体结构，阐明其获得人源受体结合能力的分子机制。本研究发现了在不含有Q226L突变的情况下，能使H5亚型HA蛋白获得人源受体结合能力的2个氨基酸突变，为监测和预警H5亚型流感病毒在人际间的流行提供了新的须关注位点。