SARS病毒S1蛋白N端片段在大肠杆菌中的表达、纯化与鉴定  

Expression and purification of recombinant N-terminal protein of SARS S1 subunit expressed in E. Coli.

作　　者：杨小昂[1] 董学员[1] 李燕[1] 李云燕[1] 徐国宾[2] 冯珍如[2] 王月丹[1] 陈慰峰[1] 

机构地区：[1]北京大学基础医学院免疫学系,北京100083 [2]北京大学第一医院检验科

出　　处：《北京大学学报（医学版）》2004年第1期79-81,共3页Journal of Peking University:Health Sciences

基　　金：国家自然科学基金 ( 3 0 3 40 0 11);国家重点基础研究发展规划项目基金 ( 2 0 0 3CB5 14 10 0 );教育部SARS攻关基金资助

摘　　要：目的 :获得纯化的重组SARS病毒S1蛋白N端部分 ,研究其与机体产生针对SARS病毒免疫应答的规律和机制。方法 :将编码SARS病毒S1蛋白N端 334个氨基酸残基的基因进行克隆 ,并在原核表达系统中表达 ,获得了纯化的重组蛋白。利用SARS患者恢复期已知的SARS抗体阳性血清 ,鉴定纯化的重组S1蛋白片段。结果 :克隆表达的重组蛋白基因序列与公布的SARS病毒S1蛋白N端的基因序列相同 ,所表达的重组蛋白相对分子质量约为 6 4 0 0 0。 3份SARS患者恢复期的血清均与重组蛋白反应 ,在相对分子质量 6 4 0 0 0处形成特异性的反应条带 ,而来自SARS流行前的正常人对照血清则不能与重组蛋白反应。结论 :获得的重组蛋白与SARS病毒抗体呈现特异性的反应 ,为进一步研究SARS病毒感染免疫应答机制和制备SARS病毒的重组疫苗奠定基础。Objective: To express and purify the recombinant N terminal protein of SARS virus S1 subunit and to study its role in SARS immune response. Methods: The gene encoding N terminal 334 amino acid residuals of SARS virus S1 subunit was cloned and expressed in E. Coli. After purification, the recombinant protein was identified by anti SARS positive sera from recovered SARS patients. The sera from health donors, which were collected before the out break of SARS, were used as negative control in the study. Results: Sequencing analysis confirmed that the desired DNA sequence in recombinant plasmid was correct and had the same sequence of natural N terminal of SARS virus S1 subunit. The molecular weight of recombinant fusion protein is about 64 000. The recombinant S1 protein could react with three antibody positive samples from recovered SARS patients, which showed specific bands at 64 000, but not with the control samples according to results of western blot. Conclusion: The recombinant N terminal protein of SARS virus S1 subunit displays specific reaction with SARS antibody and may provide a good tool for further research of immune response to SARS virus.

关 键 词：SARS 冠状病毒属 病毒蛋白质 生物合成 克隆 原核表达系统 重组疫苗 大肠杆菌 

分 类 号：R373.1[医药卫生—病原生物学]