融合蛋白NusA-hRI的高效异源表达、纯化及活性分析

人核糖核酸酶抑制因子 (human ribonuclease inhibitor, hRI) 是一种能够调节核糖核酸酶活性的酸性包浆蛋白。通过构建含SUMO、IF2、GST、NusA 、MsyB、Trx和 MBP融合标签的重组表达载体,以大肠杆菌BL21(DE3)作为宿主菌进行自诱导(auto-induction,AI)表达,从而使 hRI 的表达量得以提升。利用MagNi磁珠纯化及电泳分析hRI的表达状况,通过RNase/Sepharose亲和层析获得纯度较高的蛋白。纯化后获得的融合蛋白浓度为2 960.513mg/L,与其它公司的hRI活性进行比较,检测其酶活性约为50U/μl,并使其成功用于RNA的保护,为NusA-hRI的应用提供理论依据。

Efficient Heterologous Expression,Purification and Activity Analysis of Fusion Protein NusA-hRI

Human ribonuclease inhibitor (hRI) is an acidic pulping protein that regulates ribonuclease activity. By constructing a recombinant expression vector containing SUMO, IF2, GST, NusA, MsyB, Trx and MBP fusion tags, E.coli BL21 (DE3) was used as a host strain for auto-induction (AI) expression, thereby the expression level of hRI is improved. The expression of hRI was analyzed by MagNi magnetic bead purification and electrophoresis, and the protein with higher purity was obtained by RNase/Sepharose affinity chromatography.The concentration of the fusion protein obtained after purification was 2 960.513mg/L, which was compared with other companies, and its enzyme activity was about 50U/μl, which was successfully used for RNA protection. Purely provide a theoretic-al basis for the application of NusA-hRI.