HPV16 L2肽段偶联AP205 VLP的原核表达及免疫原性分析

中国生物工程杂志

2012, Vol. 32

Issue (6): 1-6

研究报告

HPV16 L2肽段偶联AP205 VLP的原核表达及免疫原性分析

姜博, 毕研伟, 张营营, 蔡路奎, 姬秋彦, 李智华, 徐维明

中国医学科学院/北京协和医学院医学生物学研究所昆明 650118

Prokaryotic Expression and Immunogenicity Analysis of HPV16 L2 Peptide Coupled to AP205VLP

JIANG Bo, BI Yan-wei, ZHANG Ying-ying, CAI Lu-kui, JI Qiu-yan, LI Zhi-hua, XU Wei-ming

Institute of Medical Biology, Chinese Academy of Medical Sciences, Peking Union Medical College, Kunming 650118, China

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摘要： 目的:将编码HPV16衣壳蛋白L2的65~71、112~120免疫优势表位连接到RNA噬菌体衣壳蛋白AP205氮端,组装形成病毒样颗粒,通过在大肠杆菌中实现表达及纯化,对其免疫原性进行研究。方法:合成编码AP205衣壳蛋白基因和HPV16 L2的65~71、112~120位氨基酸表位的基因序列,PCR连接并克隆至pET30a(+)原核表达载体,构建重组表达质粒pET30-AP205-HPV16ΔL2,转化大肠杆菌BL21(DH3)感受态细胞,IPTG诱导表达。表达蛋白经凝胶层析纯化及SDS-PAGE、Western blot等理化性质检测,免疫接种ICR小鼠,通过间接ELISA法检测其免疫原性。结果:成功构建重组表达质粒,重组蛋白在大肠杆菌中以可溶性表达,透射电镜观察可见典型病毒样颗粒,该VLP在动物实验中表现出较好的免疫原性。结论:成功将HPV16 L2表位偶联AP205以形成VLP,在大肠杆菌中实现可溶性表达。

关键词： 人乳头瘤状病毒; L2蛋白; AP205

Abstract: Objective: To connect the residues 65~71 and 112~120 of HPV minor capsid protein L2 to the coat protein of RNA bacteriophage AP205. The fusion protein was self-assembled into virus-like-particles (VLPs) after expressing in E.coli,then the protein was purified and the immunogenicity was analyzed. Methods: After being synthesized artificially,the AP205 coat protein gene and HPV16L2 gene were connected by PCR and cloned into vector pET-30a(+).The constructed plasmid pET30-AP205-HPV16ΔL2 was transformed to E.coli BL21(DH3), the recombinant protein was expressed under induction of IPTG and purified by Gel chromatography, then identified through SDS-PAGE and Western blot. The immunogenicity of purified product was measured by indirect ELISA after ICR mouse was immunized. Results: Recombinant plasmid pET30-AP205-HPV16ΔL2 was constructed correctly and the protein was expressed solubly in E.coli BL21(DH3). The typical VLPs can be observed through Transmission electron microscopy which Showed good immunogenicity in animal experiments. Conclusion: The HPV16L2 epitope was fused successfully to the N-terminus of AP205 coat protein,and The VLPs was self-assembled in E. coli.

Key words: Human papillomavirus L2 protein AP205 VLP

收稿日期: 2012-02-20 出版日期: 2012-06-25

ZTFLH:

Q786

基金资助: 云南省应用基础研究面上项目(2009ZC186M)

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引用本文:

姜博, 毕研伟, 张营营, 蔡路奎, 姬秋彦, 李智华, 徐维明. HPV16 L2肽段偶联AP205 VLP的原核表达及免疫原性分析[J]. 中国生物工程杂志, 2012, 32(6): 1-6.

JIANG Bo, BI Yan-wei, ZHANG Ying-ying, CAI Lu-kui, JI Qiu-yan, LI Zhi-hua, XU Wei-ming. Prokaryotic Expression and Immunogenicity Analysis of HPV16 L2 Peptide Coupled to AP205VLP. China Biotechnology, 2012, 32(6): 1-6.

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https://manu60.magtech.com.cn/biotech/CN/ 或 https://manu60.magtech.com.cn/biotech/CN/Y2012/V32/I6/1

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