Zhang Y, Hu H, Cai R, Feng Y, Zhu S, He Q, Tang Y, Xu M, Xu Y, Zhang X, Liu B, Liang Z
State Key Laboratory of Molecular Biology, Shanghai Institute of Biochemistry, Chinese Academy of Sciences, China.
Sci China C Life Sci. 1996 Jun;39(3):225-33.
A synthetic single-chain porcine insulin precursor (PIP) gene and an alpha-mating factor leader sequence (alpha MFL) gene obtained by the PCR method are inserted between the promoter and 3'-terminating sequence of the alcohol dehydrogenase gene ADH1 in plasmid pVT102-U to form plasmid pVT102-U/alpha MFL-PIP. The single-chain insulin precursor is expressed and secreted to the culture medium by Saccharomyces cerevisiae transformed by pVT102-U/alpha MFL-PIP. The precursor is purified and converted into human insulin by tryptic transpeptidation. The purified human insulin is fully active and can be crystallized. The overall yield of human insulin is 25 mg per liter of culture medium.
通过PCR方法获得的合成单链猪胰岛素前体（PIP）基因和α-交配因子前导序列（αMFL）基因被插入质粒pVT102-U中乙醇脱氢酶基因ADH1的启动子和3'-终止序列之间，形成质粒pVT102-U/αMFL-PIP。单链胰岛素前体由用pVT102-U/αMFL-PIP转化的酿酒酵母表达并分泌到培养基中。该前体经胰蛋白酶转肽作用纯化并转化为人胰岛素。纯化的人胰岛素具有完全活性且可结晶。人胰岛素的总产率为每升培养基25毫克。
