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学科主题: 基础医学
题名:
Prokaryotic expression, refolding, and purification of fragment 450-650 of the spike protein of SARS-coronavirus
作者: Zhao, JC; Zhao, ZD; Wang, W; Gao, XM
关键词: SARS-coronavirus ; spike protein ; on-column refolding
刊名: PROTEIN EXPRESSION AND PURIFICATION
发表日期: 2005-02-01
DOI: 10.1016/j.pep.2004.10.004
卷: 39, 期:2, 页:169-174
收录类别: SCI
文章类型: Article
WOS标题词: Science & Technology
类目[WOS]: Biochemical Research Methods ; Biochemistry & Molecular Biology ; Biotechnology & Applied Microbiology
研究领域[WOS]: Biochemistry & Molecular Biology ; Biotechnology & Applied Microbiology
关键词[WOS]: ACUTE RESPIRATORY SYNDROME ; ANGIOTENSIN-CONVERTING ENZYME-2 ; HIGH-LEVEL EXPRESSION ; ESCHERICHIA-COLI ; RIBONUCLEASE-A ; RECEPTOR ; STABILIZATION ; GLYCOPROTEIN ; TAG ; S1
英文摘要:

The spike (S) glycoprotein is one of the major structure proteins of SARS-associated coronavirus (CoV). Fragment 450-650 (S450-650) of the S protein contains receptor-binding domain and neutralizing epitopes. In this study. S450-650 was expresssed with a histidine tag in Escherichia coli BL21. Bacterial inclusion bodies containing the recombinant S450-650 were solubilized with 8 M urea and then applied onto a Ni-nitrilotriacetic acid column. On-column refolding and purification was performed. Reduced glutathione, and oxidized glutathione were included in the refolding buffer. In the wash and elution buffers. glycerol and glucose were necessary. T additives to prevent protein aggregation during purification. This refolding and purification procedure allowed production of S450-650 at up to 500 mug/ml in soluble form, which maintained appropriate antigenicity and immunogenicity. It was able to induce strong IgG responses in BALB/c mice. In Western blot assays, the recombinant S450-650 was recognized by monoclonal Ab against the His-tag and also sera from a convalescent SARS patient. S450-650-based ELISA system was able to detect anti-SARS-CoV IgG Abs in patient sera. (C) 2004 Elsevier Inc. All rights reserved.

语种: 英语
WOS记录号: WOS:000226437000006
Citation statistics:
内容类型: 期刊论文
版本: 出版稿
URI标识: http://ir.bjmu.edu.cn/handle/400002259/58859
Appears in Collections:基础医学院_免疫学系_期刊论文

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作者单位: Peking Univ, Ctr Hlth Sci, Dept Immunol, Sch Basic Med Sci, Beijing 100871, Peoples R China

Recommended Citation:
Zhao, JC,Zhao, ZD,Wang, W,et al. Prokaryotic expression, refolding, and purification of fragment 450-650 of the spike protein of SARS-coronavirus[J]. PROTEIN EXPRESSION AND PURIFICATION,2005,39(2):169-174.
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