胸腺素α1二聚体蛋白的克隆、表达、纯化及其生物学活性检测

Cloning,expression,purification and biologic characteristics of Tα1② protein

目的:获得具有生物学活性的重组人胸腺素α1二聚体蛋白.方法:人工合成胸腺素α1二聚体基因,并将该基因克隆入原核表达载体pET-22b(+)中.转化宿主细胞大肠杆菌BL21(DE3),经IPTG诱导表达重组Tα1②蛋白.对表达产物进行SDS-PAGE电泳和Western Blot检测分析以及生物学活性检测.结果:Tα1②Mr约为6.3×103,与理论值一致.成功纯化了Tα1②蛋白,该蛋白具有与T1抗体特异性的结合能力并能刺激小鼠T淋巴细胞增殖.结论:成功地克隆、表达和纯化Tα1②蛋白;重组Tα1②具有与化学合成Tα1相同的功能并有更高的生物学活性.

AIM：To clone,express and purify recombinant Thymosin alpha 1 ② protein （Tα1②） in E.coli. METHODS：A 189 bp recombinant gene of 2×Tα1 tandem repeats was synthesized and cloned into pET-22b（＋） vector. The expression plasmid was then transformed into E.coli. BL21（DE3）. The expression of the recombinant protein Tα1② was induced with IPTG and detected by SDS-PAGE and Western blot. The biological activity of Tα1② was identified by MTT assay. RESULTS：A novel protein with expected molecular mass was expressed. The expressed product could be recognized by a monoclonal antibody against the human Tα1. The purified Tα1② stimulated the proliferation of mice splenic lymphocytes. CONCLUSION：The recombinant Tα1② is successfully cloned,expressed and purified. The purified Tα1②protein has the same function as synthesized Tα1 but higher bioactivity.