口蹄疫病毒VP1蛋白在酵母中的表达及免疫原性分析  

Expression of FMDV VP1 protein in Pichia pastoris and its immunological activity in mice

作　　者：金华利[1] 张富春 单文娟[1] 张爱莲[1] 李轶杰[1] 王宾[1] 

机构地区：[1]新疆大学生命科学与技术学院新疆生物资源基因工程重点实验室,新疆乌鲁木齐830046

出　　处：《细胞与分子免疫学杂志》2004年第5期513-516,共4页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家高技术研究发展计划 (863)项目资助 (No .2 0 0 1AA2 1 31 1 1 );国家自然科学基金项目资助 (No.30 2 71 2 2 0 )

摘　　要：目的 :利用毕赤酵母表达系统表达牛O型口蹄疫外壳蛋白(FMDVVP1) ,并对表达的蛋白进行免疫原性鉴定。方法 :将FMDVvp1基因克隆到毕赤酵母Pichiapastoris分泌性表达载体pSuperY中 ,构建重组表达载体pSuperY/vp1,经测序证明vp1基因序列的正确性。将纯化的重组质粒经线性化酶切后 ,用电转化法将pSuperY/vp1导入毕赤酵母菌种SMD116 8H中。对表达产物用SDS PAGE和Westernblot进行分析 ,并用酵母表达的FMDVVP1蛋白免疫小鼠。结果 :以重组质粒pSuperY/vp1转化毕赤酵母菌后 ,能表达相对分子量 (Mr)为 6 6 0 0 0和4 30 0 0的FMDVVP1蛋白。动物免疫结果表明 ,FMDVVP1蛋白能诱导小鼠产生特异性的体液和细胞免疫应答。结论 :在毕赤酵母中成功地表达FMDVVP1蛋白 。AIM: To express and identify bovine O type foot and mouth disease virus protein (FMDVVP1) in yeast Pichia pastoris. METHODS: FMDV vp1 gene was cloned into secretory Pichia pastoris expression vector-pSuperY. After being linearized with enzyme digestion, the vector was transformed into Pichia pastoris SMD1168H by electroporation. The transformant was screened by zeocin. Expressed proteins in yeast were analyzed by SDS-PAGE and Western blot and then were used to immunize mice. RESULTS: The results of SDS-PAGE and Western blot demonstrated that the culture supernatant of recombinant yeast contained VP1 protein. The recombinant VP1 protein could elicit similar humoral and cellular immune responses in mice to traditional FMDV killed vaccine. CONCLUSION: FMDVVP1 is expressed successfully in yeast Pichia pastoris, which lays the foundation for further FMDV vaccine research.

关 键 词：蛋白 酵母表达 VP1 免疫原性 毕赤酵母菌 重组质粒 细胞免疫应答 口蹄疫病毒 VP1基因 FMDV 

分 类 号：R373[医药卫生—病原生物学] S852[医药卫生—基础医学]