O型口蹄疫病毒VP1表位重组蛋白疫苗的构建、表达和纯化

目的 为了克服灭活口蹄疫病毒疫苗可能存在的传播病毒的潜在危险，构建一种能预防O型口蹄疫病毒感染的VPI表位重组蛋白疫苗。方法采用O型口蹄疫病毒（FMDV）表面VP1蛋白上的B细胞表位肽和T细胞表位肽，以PCR扩增和克隆连接等方法构建VP1表位六聚体重组蛋白（vP1epi）基因，并在大肠杆菌中进行诱导表达，镍亲和层析纯化。用ELISA法检测VPlepi免疫豚鼠血清中抗FMDV抗体的水平。结果构建了一种由FMDV表面vP1中B细胞表位肽重复六次的蛋白质多肽，采用pET28原核表达系统，在大肠杆菌BL21（DE3）中获得了高效表达，表达产量约占菌体蛋白的30％左右，经镍亲和层析后获得纯度高于90％的VP1表位重组蛋白。VP1epi免疫的豚鼠血清中含有一定量的抗FMDV抗体。结论VP1epi重组蛋白能诱导机体产生抗O型FMDV的抗体，说明这种重组蛋白可能成为预防O型FMDV感染的基因工程蛋白质疫苗。

Construction, expression, and purification of a recombinant epitope vaccine for type O foot-and mouth disease virus

Objective To developed a recombinant protein vaccine with epitopes on VP1 protein for prevention of foot-and-mouth disease virus(FMDV) spreading by inactive-FMD vaccine.Methods B cell epitopes and T cell epitopes on VP1 protein of type O FDMV were used to construct the gene encoding VP1 epitope-recombinant protein(VP1epi) by PCR and DNA cloning.The VP1epi protein was expressed in E.coli with pET28 prokaryotic expression system and purified by nickel affinity chromatography.The anti-FMDV antibody in sera of guinea pig immunized with VP1epi protein was detected by ELISA method.Results VP1epi protein constructed by epitopes on VP1 protein of type O FMDV was highly expressed in BL21(DE3) and the expression yield was about 30% of total bacterial protein.The purity of VP1epi protein purified by nickel affinity chromatography was more than 90%.The anti-FMDV antibody was found in sera of VP1epi-immunized guinea pig.Conclusion The recombinant VP1epi protein can induce animal to produce specific anti-FMDV antibody and may be used as a novel genetic protein vaccine for prevention of type O FMDV infection.