展示日本乙型脑炎病毒B细胞表位和T细胞表位的猪细小病毒病毒样颗粒的制备

Construction of porcine parvovirus-like particles harboring B cell and T cell epitopes of Japanese encephalitis virus

为增强日本乙型脑炎病毒表位疫苗的免疫原性,提高其免疫效果,将日本乙型脑炎病毒(JEV)E蛋白中编码的B细胞表位(150～156aa,307～316aa,327～333aa,386～399aa)及T细胞表位(60～68aa,436～445aa)序列连接于猪细小病毒(PPV)VP2的5′端,并插入到原核表达质粒pCold-Ⅰ中,构建成重组质粒pMEP-VP2,转化大肠杆菌BL21(DE3),使重组蛋白MEP-VP2得到表达。Western-blot分析表明,表达产物能被抗JEV多抗和抗PPV多抗识别。电镜观察显示,表达的重组蛋白MEP-VP2能够在体外自我组装成病毒样颗粒PPV-VLP(JEV)。结果表明,N端连接JEV的多表位肽的猪细小病毒的VP2蛋白能体外组装成有活性的病毒样颗粒。

To improve the immunogenicity of Japanese encephalitis virus（JEV） epitope vaccine, the gene fragments encoding B-cell epitopes（aa 150--156, aa 307--316,aa 386--399） and T cell epitopes（aa 60 --68,aa 436--445） of JEV were assembled with VP2 gene of porcine parvovirus（PPV） and inserted into plasmid pCold- I to construct recombinant plasmid pMEP-VP21. The resulted plasmid was transformed into E. coli BL21（DE3） to express the recombinant protein. The expression product was detected by Westernblot. In result, the recombinant protein could be distinguished by anti-JEV polyclonal antibody and anti- PPV polyclonal antibody. In addition,the virus-like particles of PPV-VLP（JEV） was observed by electron microscopy.