人源FGF-21的高效可溶性表达及其调节血糖功能的初步研究

Efficient expression of soluble human FGF-21 and its glucose regulation activity

将人源FGF-21基因亚克隆至pSUMO表达载体上,在大肠杆菌Rosetta(DE3)中诱导表达,在pSUMO表达体系中的重组人源FGF-21以可溶形式表达,重组蛋白经镍离子螯合柱(Ni-NTA)纯化,透析后利用SUMO蛋白酶I切割融合蛋白,获得纯度较高的重组人源FGF-21。将3T3-L1成纤维细胞分化成脂肪细胞,经微量化的GOD-POD法检测培养基中葡萄糖含量,统计学分析葡萄糖消耗率。与未经处理的脂肪细胞对照组相比,经重组人源FGF-21处理后脂肪细胞对葡萄糖的摄取利用显著增加,残存在培养基中的葡萄糖含量明显减少(P<0.05,P<0.001)。

The cDNA of human FGF-21 was subcloned into the pSUMO expression vector and the fusion protein was induced to express in Escherichia coli Rosetta （DE3）. The recombinant hFGF-21 was expressed in soluble form in the pSUMO expression system. The recombinant fusion protein was purified by Ni-NTA column. The purified recombinant protein was dialyzed against PBS for reature. To obtain pure and active recombinant protein, the fusion protein was subjected to cleavage with SUMO protease I. To examine glucose regulation activity of hFGF-21, 3T3-L1 pre-adipocytes were differentiated into adipocytes, glucose up-take activity of hFGF-21 was examined by glucose oxidase and peroxidase （GOD-POD） assay. Compared with no stimulation control, the recombinant hFGF-21 treatment led to a significant increase in glucose consumption of adipocytes and a significant decrease in concentration of glucose in the medium （ P 〈 0.05, P 〈 0.001）.