苍白密螺旋体Tp0751黏附蛋白的表达、纯化及免疫反应性分析

Expression and Purification of Tp0751 Recombinant Protein of Treponema pallidum and Its Immunoresponse Analysis

目的表达苍白密螺旋体(梅毒螺旋体)黏附蛋白Tp0751,纯化表达产物并进行免疫反应性分析,为探索Tp0751重组蛋白在梅毒致病过程中的作用奠定基础。方法通过生物信息学分析,去除Tp0751信号肽序列,构建原核表达体进行诱导表达;Ni亲合层析柱纯化重组蛋白,Western-印迹检测其免疫反应性。结果成功构建了PET-28a(+)-0751原核表达载体,经表达、纯化后获得了相对分子量约为26×103的融合蛋白,其表达产物占全菌总蛋白的>30%,并且主要可溶性形式存在;Western-印迹检测其能与梅毒患者阳性血清发生特异性反应,具有良好的抗原性。结论重组表达的Tp0751黏附蛋白为可溶性蛋白,且具有良好的免疫反应性,为进一步研究其在Tp0751黏附蛋白在梅毒致病过程中的作用和其生物学功能奠定了基础。

OBJECTIVE To express Tp0751 laminin-binding adhesin of Treponema pallidum, and assess the immunoresponse. METHODS The Tp0751 ORF without upstream non-coding region was ligased into the expression vector PET-28a（＋）, and expressed in E. coli R2566. Its immunogen was analyzed by Western-blot and ELISA. RESULTS A fusion protein with relative molecular mass about 26 × 10^3 was attained after expression and purification. Western blot proved that the recombinant protein can specifically react with T. pallidurn IgG positive sera. CONCLUSIONS The expressed recombinant protein shows excellent immunoresponse, and the results lay the foundation for the research on its function to T. pallidum infection.