布氏杆菌PBP_(39)蛋白抗原表达及免疫效果的研究

Expression and immunogenicity of PBP_(39) recombinant protein of Brucella

目的 获得纯化的布氏杆菌PBP3 9重组蛋白抗原,观察PBP3 9重组蛋白的免疫原性。方法 扩增不同种布氏杆菌PBP3 9基因,测序、连接表达载体PET3 2a ,诱导表达,柱纯化PBP3 9重组蛋白抗原,免疫BALB/c小鼠,ELISA检测抗体及抗体亚型。结果 我国牛、羊、猪布氏杆菌PBP3 9基因与国外已报道的PBP3 9编码基因不完全相同。在蛋白N端5 8位碱基后多一G ,造成移码突变。故在85、86、87位编码终止子TCA。而在13 4、13 5、13 6位的ATG又编码了新的起始密码。布氏杆菌PBP3 9蛋白在大肠杆菌高效表达,表达量达40 % ,纯化重组蛋白抗原免疫BALB/c小鼠,产生了较高水平的特异性抗体免疫应答,抗体亚型主要是IgG1。结论 我国牛、羊、猪布氏杆菌PBP3 9编码抗原蛋白同国外已报道的不完全相同,纯化的PBP3 9重组蛋白抗原可诱导高水平的抗体反应,为筛选布氏杆菌病新型疫苗保护性抗原分子奠定了基础。

Objective To obtain recombinant PBP 39 protein of Brucella and study its immunity. Methods The PBP 39 protein DNA was amplified by PCR, cloned into PET32a vector and transformed into E.coli BL21 for expression. The BALB/c mice were immunized with the purified recombinant protein. The antibody level and specific antibody against PBP 39 protein was detected by ELISA. Results The site of ATG was not as the same as PBP 39 reported in GenBank. At the N terminal, there was a frameshift mutation at 58 site. The Brucella PBP 39 protein expressed highly in the E.coli. Experimental animals exhibited specific antibody that was detectable by ELISA. IgG1-specific antibody titers of the recombinant PBP 39 protein were higher than IgG2a-specific antibody. Conclusion The PBP 39 protein is special in China and induce immune response.