去N端信号肽的水痘-带状疱疹病毒糖蛋白E的原核可溶性表达及其免疫原性评价  

Soluble prokaryotic expression of varicella-zoster virus glycoprotein E without N-terminal signal peptide and its immunogenicity evaluation

作　　者：魏明童 夏兵兵 何志远 周炜 刘兴东 赵俊 陈敬贤[4] 王明丽[2,4] WEI Mingtong;XIA Bingbing;HE Zhiyuan;ZHOU Wei;LIU Xingdong;ZHAO Jun;CHEN Jingxian;WANG Mingli(The First Clinical Medical College of Anhui Medical University,Hefei 230032,Anhui Province,China;Wuhu Interfell Research Institute,Wuhu 241000,Anhui Province,China;Second Clinical Medical College of Anhui Medical University,Hefei 230032,Anhui Province,China;Institute of Clinical Virology,Anhui Medical University,Hefei 230000,Anhui Province,China)

机构地区：[1]安徽医科大学第一临床医学院,安徽合肥230060 [2]芜湖英特菲尔生物制品产业研究院有限公司,芜湖241000 [3]安徽医科大学第二临床医学院,合肥230060 [4]安徽医科大学临床病毒学研究所,合肥230032

出　　处：《微生物与感染》2020年第6期377-384,共8页Journal of Microbes and Infections

基　　金：安徽省高等学校省级质量工程项目(2012sjjd014)。

摘　　要：水痘-带状疱疹病毒(varicella zoster virus,VZV)糖蛋白E(glycoprotein E,gE)是VZV亚单位疫苗的主要候选蛋白,但目前原核表达系统制备的gE蛋白以包涵体形式为主,可溶性差。本研究采用去除第1~30氨基酸序列的VZV gE胞外域基因,将其与原核表达载体pET32a连接,并转化至感受态细胞BL21(DE3)中。使用异丙基-β-D-硫代半乳糖苷(Isopropylβ-D-thiogalactoside,IPTG)诱导表达,His-tag柱纯化重组gE蛋白,蛋白质印迹法(Western blot,WB)检测其特异性。用该重组gE蛋白免疫BALB/c小鼠制备多克隆抗体,酶联免疫吸附试验(enzyme linked immunosorbent assay,ELISA)和间接免疫荧光法检测多克隆抗体效价及特异性。结果显示,BL21/pET32a-VZV gE工程菌可以表达可溶性重组gE蛋白,纯化后纯度约为90%。WB鉴定该重组蛋白具有良好的免疫反应性。ELISA检测显示小鼠抗VZV gE多克隆抗体效价>1∶10000,间接免疫荧光实验结果显示该抗体特异性较高。结果表明,本研究在原核表达系统中成功表达可溶性重组VZV gE蛋白,同时该蛋白具有较强的免疫原性,这为VZV gE亚单位疫苗的研制和大规模生产奠定了基础。Varicella zoster virus(VZV)glycoprotein E(gE)is the main candidate protein for VZV subunit vaccine.Unfortunately,the expression of gE in prokaryotic expression was mainly found in the inclusion body.In this study,a deletion of N-terminal 30 amino acids of gE was constructed on pET32a,expressed in BL21(DE3).The soluble gE was purified and used to make polyclonal antibodies in BALB/c mice.The titer and specificity of polyclonal antibodies were determined by enzyme linked immunosorbent assay(ELISA)and indirect immunofluorescence.The results showed that the soluble recombinant gE produced by BL21/pET32a-VZV gE system is a practical way for vaccine development.

关 键 词：水痘-带状疱疹病毒 糖蛋白E 原核表达 多克隆抗体 免疫评价 

分 类 号：R373.12[医药卫生—病原生物学]