Kobayashi M, Sugibayashi M, Sasaoka T, Egawa K, Shigeta Y, Tamaki M, Nakamura E, Teraoka H
Third Department of Medicine, Shiga University of Medical Science, Ohtsu, Japan.
Biochem Biophys Res Commun. 1990 Mar 30;167(3):1073-8. doi: 10.1016/0006-291x(90)90632-w.
To study whether the G----T point mutation of insulin proreceptors at the cleavage site which changed -Arg-Lys-Arg-Arg- to -Arg-Lys-Arg-Ser- caused unprocessed insulin receptors with decreased insulin binding affinity, we performed transfection of cDNA with the mutation in COS 7 cells and examined the expressed insulin receptors. After site-directed mutagenesis, an expression vector pGEM3SV was used to make a plasmid which contained full-length HIRcDNA behind SV40 early promoter. Transfection of normal HIR cDNA produced normal insulin receptors on the plasma membranes in COS 7 cells. However, transfection of cDNA with the mutation resulted in the presence of 210K proreceptors in the plasma membranes with decreased insulin binding ability (35% of normal). These results suggest that the mutation, not the defect of converting enzyme, was the cause for unprocessed insulin proreceptors in the patients with insulin resistance.
为研究胰岛素原受体在裂解位点的G----T点突变（该突变将-Arg-Lys-Arg-Arg-变为-Arg-Lys-Arg-Ser-）是否导致具有降低的胰岛素结合亲和力的未加工胰岛素受体，我们在COS 7细胞中进行了携带该突变的cDNA转染，并检测了表达的胰岛素受体。经过定点诱变后，使用表达载体pGEM3SV构建了一个质粒，该质粒在SV40早期启动子后含有全长HIR cDNA。正常HIR cDNA的转染在COS 7细胞的质膜上产生了正常的胰岛素受体。然而，携带该突变的cDNA转染导致质膜中出现210K原受体，其胰岛素结合能力降低（为正常的35%）。这些结果表明，该突变而非转化酶缺陷是胰岛素抵抗患者中未加工胰岛素原受体的原因。
