重组大肠杆菌肠毒素蛋白LTA_(192)-STa_(13)的表达及其诱导小鼠体液免疫的研究  

Expression of recombinant enterotoxin protein LTA_(192_-STa_(13) in Escherichia coli and the humoral immune response induced in mice

作　　者：杜元策[1] 沙木哈吾.别开 刘文鑫[1] 关玮琨[1] 孟相秋[1] 韩林林[1] 李金萍[1] 袁超文[1] 唐杰[1] 李星月[1] 赵志腾 师东方[1] 

机构地区：[1]东北农业大学动物医学学院,黑龙江哈尔滨150030 [2]克孜勒希力克乡畜牧兽医站,新疆富蕴836106

出　　处：《中国预防兽医学报》2015年第4期304-309,共6页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家科技支撑计划项目(2012BAD12B03;2012BAD12B05);黑龙江省科技计划项目(GC12B303)

摘　　要：为研究重组大肠杆菌肠毒素融合蛋白LTA192-STa13的免疫原性,本研究将产肠毒素大肠杆菌(ETEC)elt A和est A基因定点突变后构建融合基因,进行重组蛋白LTAR192G-STaG13Q的表达。利用纯化的重组蛋白和灭活菌液分别免疫小鼠,比较不同抗原诱导小鼠体液免疫应答能力,以及体外免疫血清对毒素的中和作用。结果显示,各组免疫小鼠均可以产生高滴度抗LTA和STa抗体,其血清抗体可以通过乳汁传递给新生仔鼠,其中蛋白免疫组血清抗体效价更高。体内外中和试验证明,实验组小鼠三免后的血清能够中和LT和STa毒素对实验鼠的毒性作用,并且能够中和LT对Y-1细胞的毒性作用,蛋白免疫组血清抗体中和效价可达到1∶89.13,明显高于灭活菌免疫组。研究表明,重组蛋白LTA192-STa13对小鼠具有良好的免疫原性,能够诱导免疫鼠产生良好的体液免疫应答,可以作为ETEC亚单位疫苗的候选抗原。To study the immunogenicity of fusion enterotoxin protein LTA192-STa13 of E.coli, the eltA-stA fusion gene was constructed by SOE-PCR following the mutation of eltA and expressed in E.coli. The mice were immunized with recombinant estA genes, and the recombinant protein LTAR192G-STaG13Q was proteins and inactivated recombinant E.coli, and the level of the humoral immune response induced by different antigens and neutralizing effects of the immune serum against toxin were compared. The results showed that the high titers of anti-LTA and anti-STa antibodies in serum were induced which was able to transfer to offspring via foremilk. The serum antibody titer of the group immunized with fusion protein was higher than that of the group immunized with inactivated E.coli. The serum antibodies after the third immunization not only neutralize native LT and STa toxin, but also neutralize toxic effects of LT to Y-1 cells. The neutralization titers of serum antibodies against LT of mice immunized with fusion protein LTA_192-STa_13 was 1：89.13, higher than that of mice immunized with inactivated E.coli. The results demon- strated that the fusion protein LTA192-STa13 possessed a strongle immunogenicity, and induced an efficient humoral immune response in mice, which could be candidate antigen for ETEC subunit vaccine.

关 键 词：大肠杆菌 肠毒素 重组蛋白 体液免疫 

分 类 号：S852.61[农业科学—基础兽医学]