摘要
本研究旨在制备人GⅡ.14诺如病毒的(Norovirus,NoV)病毒样颗粒(Virus⁃like particle,VLP),探索其抗原性与组织血型抗原结合的特征。利用杆状病毒系统表达人GⅡ.14 NoV VP1蛋白,通过超速离心和分子筛层析纯化VLP;将VLP免疫动物制备兔多抗血清,通过免疫印迹(Western blot,WB)、酶联免疫吸附(ELISA)进行抗原性评价及多抗功能验证;利用唾液结合实验检测人GⅡ.14 VLP的糖结合特征;通过蛋白序列及结构比较分析GⅡ.14 P蛋白潜在的糖结合位。纯化获得人GⅡ.14 VP1蛋白,电镜观察显示可以形成结构较为均一的VLP;制备的兔多抗能与GⅡ.14 VLP特异性结合,与检测的其他基因型VLP没有交叉反应;唾液结合实验表明GⅡ.14 VLP与人A/B/O/AB型别唾液都有较好的结合;序列和结构分析显示GⅡ.14 P蛋白模拟结构与GⅡ.9最为接近,具有与GⅡ.9以及流行株GⅡ.4等相似的潜在糖结合区。本研究表明GⅡ.14具有较为广泛的唾液结合特性,通过序列比对和结构学方法分析了GⅡ.14潜在的受体结合机制,为不同型别NoVs的流行监测提供了更多依据。
This study aimed to get the virus⁃like particles(VLPs)of human GII.14 norovirus(NoV)and explore its binding characteristics with histo⁃blood group antigens(HBGAs).The VP1 protein of human GII.14 NoV was expreesed by baculovirus system.VLPs were purified by ultracentrifugation and gel⁃filtration chromatography.Rabbit polyclonal antisera were generated by immunizing animals with GII.14 VLPs.The function of polyclonal antibodies was verified by western blotting and enzyme⁃linked immunosorbent assay.The saliva⁃binding assay was carried out to detect the glycan⁃binding characteristics of human GII.14 VLPs.Potential glycan binding sites of the GII.14 P protein were analyzed by sequence alignment and structural comparison.The purified GII.14 VP1 protein formed relatively uniform VLPs when observed by electron microscopy.Rabbit polyclonal antibodies could specifically bind to GII.14 VLPs with no obvious cross⁃reactivity with other tested VLPs.GII.14 VLPs bound well to human saliva of A/B/O/AB types according to the saliva⁃binding assay.Sequence and structural analyses showed that the simulated structure of the GII.14 P protein was closest to GII.9 P protein,and possessed similar potential glycan⁃binding sites to that of GII.9 and the epidemic GII.4.Our data indicate that GII.14 has relatively broad binding characteristics to saliva.The potential glycan⁃binding mechanism of GII.14 was illustrated through sequence alignment and structural comparison.Our results provide more evidence for the epidemic monitoring of NoVs of different genotypes.
作者
欧阳瑄泽
柴鹏弟
宋敬东
靳淼
李利利
李金松
孙晓曼
段招军
OUYANG Xuanze;CHAI Pengdi;SONG Jingdong;JIN Miao;LI Lili;LI Jinsong;SUN Xiaoman;DUAN Zhaojun(School of Public Health,Gansu University of Chinese Medicine,Lanzhou 730000,China;National Key Laboratory of Intelligent Tracking and Forecasting for Infectious Diseases(NITFID),NHC Key Laboratory for Medical Virology and Viral Diseases,National Institute for Viral Disease Control and Prevention,Chinese Center for Disease Control and Prevention,Beijing 102206,China)
出处
《病毒学报》
CAS
CSCD
北大核心
2024年第4期705-715,共11页
Chinese Journal of Virology
基金
北京市自然科学基金(项目号:L232011),题目:我国诺如病毒基因变异变迁及所致胃肠炎流行特征和经济负担研究。
关键词
诺如病毒
病毒样颗粒
组织血型抗原
Norovirus
Virus⁃like particle(VLP)
Histo⁃blood group antigen(HBGA)
作者简介
欧阳瑄泽(2000⁃),女,Email:1820425218@qq.com;通信作者:段招军(1973⁃),男,研究员,从事病毒性腹泻预防控制相关研究,Email:zhaojund@126.com。
