梅毒螺旋体 Tp0844重组蛋白的表达、纯化及免疫反应性研究  

Expression, purification and evaluation of immunoreactivity of the recombinant protein Tp0844 of Treponema pallidum

作　　者：刘双全[1] 刘琼[1] 

机构地区：[1]南华大学附属第一医院检验科,湖南衡阳421001

出　　处：《中华皮肤科杂志》2015年第5期326-328,共3页Chinese Journal of Dermatology

基　　金：国家自然科学基金（81201331）;湖南省自然科学基金（11JJ4076）;湖南省教育厅青年基金（118107）

摘　　要：目的：克隆、表达梅毒螺旋体重组蛋白 Tp0844，纯化表达产物并进行免疫反应性分析，筛选与宿主具有高反应性的 Tp 主要蛋白。方法通过生物信息学分析，获取 Tp0844基因序列，构建原核载体进行诱导表达；Ni-NTA 亲合层析柱纯化重组蛋白，Western 印迹法检测其重组蛋白与梅毒阴阳性血清的反应情况。结果成功构建 PET-30a（＋）-Tp0844原核表达重组体，经诱导表达后发现该重组体可高表达出可溶性的重组蛋白，经亲和层析纯化后获得了相对分子质量为43000的重组蛋白；以梅毒 IgG 抗体阴、阳性血清为一抗，采用 Western 印迹法检测发现，Tp0844重组蛋白与梅毒阳性血清能发生明显特异反应，而与健康阴性血清未出现反应条带。结论可溶性重组表达的 Tp0844蛋白具有较好的免疫反应性，可作为梅毒发病机制研究的候选抗原。Objective To clone, express, purify and evaluate the immunoreactivity of the recombinant protein TP〈0844 of Treponema pallidum （Tp）, and to screen major Tp proteins with high host reactivity. Methods The TP〈0844 gene sequence was obtained through bioinformatics analysis. A prokaryotic expression vector of the TP〈0844 gene was constructed and transformed into E. coli BL21 followed by isopropyl-1-thio-β-D-galactopyranoside （IPTG）induction for the expression of the recombinant protein TP〈0844. Nickel-NTA affinity chromatography columns were utilized to purify the recombinant protein, and Western blotting was performed to evaluate the reactivity of the recombinant protein with sera positive or negative for anti-Tp IgG antibodies. Results The recombinant prokaryotic expression vector PET-30a （＋）-TP〈0844 was successfully constructed. After IPTG induction, a soluble recombinant protein with a relative molecular mass of about 43 000 was highly expressed, and purified by affinity chromatography. Western blotting showed that the TP〈0844 recombinant protein specifically reacted with anti-Tp IgG antibody-positive sera, but not with anti-Tp IgG antibody-negative sera. Conclusions The soluble recombinant protein TP〈0844 has good immunoreactivity, and can serve as a candidate antigen for investigation into the pathogenesis of syphilis.

关 键 词：苍白密螺旋体 重组蛋白质类 TP0844 免疫学 免疫反应性 

分 类 号：R392[医药卫生—免疫学]