九种冠状病毒核衣壳蛋白的原核表达及纯化

基础医学与临床 ›› 2009, Vol. 29 ›› Issue (5): 499-503.

九种冠状病毒核衣壳蛋白的原核表达及纯化

许慧雯王彦斌吴超王健伟洪涛

中国医学科学院北京协和医学院病原生物学研究所中国医学科学院北京协和医学院病原生物学研究所

收稿日期:2009-03-02 修回日期:2009-03-13 出版日期:2009-05-25 发布日期:2009-05-25
通讯作者: 王健伟

Expression and purification of nine nucleocapsid proteins from coronaviruses

Hui-wen XU, Yan-bin WANG, Chao WU, Jian-wei WANG, Tao HONG

Institute of Pathogen Biology，CAMS& PUMC Institute of Pathogen Biology，CAMS& PUMC

Received:2009-03-02 Revised:2009-03-13 Online:2009-05-25 Published:2009-05-25
Contact: Jian-wei WANG,

摘要/Abstract

摘要： 目的探索SARS冠状病毒（SARS-CoV）、HCoV-229E、HCoV-OC43、HCoV-NL63、HCoV-HKU1等5种人类冠状病毒以及牛冠状病毒（BCoV）、鼠肝炎病毒（MHV）、猫传染性腹膜炎病毒（FIPV）和禽传染性支气管炎病毒（IBV）4种动物冠状病毒核衣壳（N）蛋白在大肠杆菌中克隆表达及纯化条件。方法利用原核表达载体pET30a（+）经IPTG诱导表达重组蛋白，并采用离子交换及镍离子螯合亲和层析法对重组蛋白进行纯化，用SDS-PAGE及Western blot对重组蛋白进行鉴定。结果成功表达、纯化出9种冠状病毒N蛋白，纯度达90%以上。结论在原核系统中表达得到了高纯度的9种冠状病毒N蛋白，为相关功能性研究奠定了基础。

关键词: 冠状病毒, 核衣壳蛋白, 纯化

Abstract: Objective To explore the conditions of the expression and purification of nucleocapsid proteins of human coronaviruses SARS-CoV, HCoV-229E, HCoV-OC43, HCoV-NL63 and HCoV-HKU1, animal coronaviruses bovine coronavirus, murine hepatitis virus, feline infectious peritonitis virus, and infectious bronchitis virus. Methods The pET-30a-based recombinant plasmids containing full length nucleocapsid(N) protein of coronaviruses were transformed into E.coil BL21(DE3) competent cells and were induced to express N proteins by IPTG. The expression products were purified by ion exchange chromatography and Ni 2+ affinity chromatography, and were verified by SDS-PAGE and Western blot. Results Nine coronavirus nucleocapsid proteins with correct Mr were solubly expressed and highly purified with purity >90%. Conclusion Successfully expressed nine recombinant nucleocapsid proteins with high purity in E. Coli，which provides materials to study the function of these coronavirus N proteins.

Key words: SARS-CoV, Nucleocapsid protein, Purification

许慧雯王彦斌吴超王健伟洪涛. 九种冠状病毒核衣壳蛋白的原核表达及纯化[J]. 基础医学与临床, 2009, 29(5): 499-503.

Hui-wen XU; Yan-bin WANG; Chao WU; Jian-wei WANG; Tao HONG. Expression and purification of nine nucleocapsid proteins from coronaviruses[J]. Basic & Clinical Medicine, 2009, 29(5): 499-503.

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