钩端螺旋体多抗原肽的构建及免疫性分析

目的 构建问号钩端螺旋体(简称钩体)主要外膜蛋白OmpL1、LipL21和LipL32优势抗原表位的串联基因及其表达系统,了解该重组蛋白的免疫活性.方法 采用噬菌体M13KE表面展示技术结合Western blot分析,鉴定了OmpLl、LipL21和LipL32的优势抗原表位,人工合成优势抗原表位串联基因并构建其原核表达系统.SDS-PAGE检测重组蛋白的表达情况;Western blot及ELISA鉴定重组蛋白的免疫活性.结果 该合成基因在原核表达系统中得到了有效表达,且表达产物主要以可溶性形式存在.Western blot和ELISA结果 显示该重组蛋白能与兔抗钩体全菌抗体及不同血清群的钩体病人血清中的抗体产生免疫反应.结论 本研究成功构建了钩体多表位串联基因及其表达系统,所表达目的 蛋白具有良好的免疫活性,且对不同血清群型抗体之间均有免疫原活性.

Construction of multiple antigenic peptide and the immunity analysis

Objective To construct the multiple antigenic peptide (MAP) gene and E. coli ex-pression system, based on the out membrane protein OmpL1, LipL21 and LipL32 from Leptospira interro-gans, and better understanding of the immunological activity of the recombinant protein. Methods Using MI3KE display and Western blot, the advantage epitopes of OmpL1, LipL21 and LipL32 were identified and used to synthesize a new gene, then its prokaryotic expression system was constructed. The expression of re-combinant protein was determined by SDS-PAGE. The immunity activity of the recombinant protein was iden-tified by Western blot and ELISA. Results The synthetic gene was effectively expressed in E. coli and mainly presented in soluble form. The expression protein could react with the antileptospirosis antibodies in rabbit and human sera, which contained different serogroups. Conclusion The recombinant MAP gene of leptospires was successfully constructedand and expressed in E. coli. The recombinant protein had a good immune activity, and could cross-reacted with antileptospirosis antibodies from different serogroups.