人类HBxAg蛋白反式激活基因5的原核表达及其蛋白纯化

Prokaryotic expression and protein purification of human XTP5 gene transactivated by X protein of hepatitis virus B

为了更深入研究XTP5蛋白(也称人类次要组织相容性抗原(mHA-8))的生物学功能,以HepG2细胞总RNA为模板、应用RT-PCR技术获得XTP5基因,再利用基因工程技术对其进行克隆和原核重组表达载体构建和诱导表达,成功克隆XTP5基因,并成功构建原核重组表达空载体pET-32a(+)-XTP5;阳性重组质粒转化大肠杆菌E.coliBL21后,在37℃经终浓度1 mmoL/L IPTG诱导5 h后,受体菌大量表达76.88 ku XTP5重组蛋白;West-ern-blot证实宿主菌表达的蛋白特异性好;肝素亲和层析柱纯化表达产物XTP5蛋白效果良好。结果表明,成功克隆了人类XTP5基因,并获得纯度较高的融合蛋白XTP5。

To study biological function of XTP5 （mHA-8） furthur, the RT-PCR and bioengineering were used to clone and express XTP5 gene. XTP5 gene （1 527 bp） and its recombinant expression vector （pET-32a （＋）-XTPS） were successfully carried out. Moreover, pET-32a （＋）-XTP5 was expressed for five hours with 1.0 mmoL/L IPTG （final concentration） at 37 ℃. The recombination protein showed iden- tical molecular weight with 76.88 ku and was confirmed by Western-blot. The purified recombinant protein was acquired by the protein purification system. Cloning and prokaryotic expression and the purified protein of XTP5 gene were successfully performed.