Wei G, Hu M H, Tang J G
Department of Biochemistry and Molecular Biology, College of Life Sciences, Peking University, Beijing, China.
Biochem Mol Biol Int. 1995 Jan;35(1):37-46.
A fusion gene encoding double-C-peptide human proinsulin was constructed by insertion of a DNA fragment encoding human C-peptide into the 5'-terminal C-peptide coding sequence of a synthetic human proinsulin gene with correct reading frame and over-expressed in E. coli. The purified double-C-peptide human proinsulin shows decreased activity in receptor binding and insulin immune assays as compared with human proinsulin. Disulphide bond reconstitution studies demonstrate that there is not much more influence of the protein concentration on the yield of refolded double-C-peptide human proinsulin. The double-C-peptide human proinsulin shows a 1.86-fold human C-peptide immune activity as compared with that of human proinsulin and gives a good yield of the molecule with correct disulphide bonds in reconstitution studies strongly suggesting the existence of very flexible conformation of the C-peptide.
通过将编码人C肽的DNA片段插入具有正确阅读框的合成人胰岛素原基因的5'-末端C肽编码序列中，构建了编码双C肽人胰岛素原的融合基因，并在大肠杆菌中过表达。与人生胰岛素原相比，纯化的双C肽人胰岛素原在受体结合和胰岛素免疫测定中显示出活性降低。二硫键重构研究表明，蛋白质浓度对复性双C肽人胰岛素原的产量影响不大。与人生胰岛素原相比，双C肽人胰岛素原显示出1.86倍的人C肽免疫活性，并且在重构研究中具有正确二硫键的分子产量良好，这强烈表明C肽存在非常灵活的构象。
