Kole H K, Liotta A S, Kole S, Roth J, Montrose-Rafizadeh C, Bernier M
Diabetes Section, Laboratory of Clinical Physiology, NIA, National Institutes of Health, Baltimore, Maryland 21224, USA.
J Biol Chem. 1996 Dec 6;271(49):31619-26. doi: 10.1074/jbc.271.49.31619.
The role of the insulin receptor COOH-terminal domain in the regulation of insulin signal transduction was explored with a variety of synthetic peptides. One of the peptides, termed peptide HC, whose structure corresponds to residues 1293-1307 of the insulin proreceptor sequence, enhanced insulin-stimulated autophosphorylation of the insulin receptor in cell-free systems and in semipermeabilized Chinese hamster ovary (CHO) cells that had been transfected with an expression plasmid encoding the human insulin receptor (CHO/HIRc) at concentrations where there was no detectable effect on basal autophosphorylation levels or on receptor dephosphorylation. A lipophilic analogue of peptide HC, stearyl peptide HC, added to intact CHO/HIRc cells enhanced significantly insulin-stimulated insulin receptor autophosphorylation while having no effect on ligand-stimulated receptor phosphorylation in CHO cells overexpressing either the IGF-1 receptor or epidermal growth factor receptor. Addition of stearyl peptide HC to CHO/HIRc cells resulted in a 2.4 +/- 0.3-fold increase in the amount of insulin-stimulated phosphatidylinositol 3-kinase detected in anti-IRS-1 immunoprecipitates and a 2.1 +/- 0.6-fold increase in the levels of tyrosine phosphorylation of mitogen-activated protein kinase in response to insulin. Finally, a derivative of peptide HC coupled to a biotin moiety was prepared and showed to bind with the beta-subunit of the wild-type insulin receptor and a truncated receptor that lacks 43 amino acids from its carboxyl terminus. However, there was little binding, if any, of the peptide with the IGF-1 receptors or the epidermal growth factor receptors. Taken together, our data demonstrate that a pentadecapeptide related to the carboxyl terminus of the insulin receptor binds to the insulin receptor beta-subunit and that this interaction may contribute to the increased receptor's intrinsic activity and signal transduction.
利用多种合成肽对胰岛素受体COOH末端结构域在胰岛素信号转导调节中的作用进行了探索。其中一种肽称为肽HC，其结构对应于胰岛素原受体序列的1293 - 1307位残基，在无细胞系统以及用编码人胰岛素受体的表达质粒转染的半透化中国仓鼠卵巢（CHO）细胞（CHO/HIRc）中，该肽能增强胰岛素刺激的胰岛素受体自身磷酸化，且在对基础自身磷酸化水平或受体去磷酸化无明显影响的浓度下即可发挥作用。将肽HC的亲脂类似物硬脂酰肽HC添加到完整的CHO/HIRc细胞中，可显著增强胰岛素刺激的胰岛素受体自身磷酸化，而对过表达IGF-1受体或表皮生长因子受体的CHO细胞中配体刺激的受体磷酸化无影响。向CHO/HIRc细胞中添加硬脂酰肽HC，在抗IRS-1免疫沉淀中检测到的胰岛素刺激的磷脂酰肌醇3激酶量增加了2.4±0.3倍，且响应胰岛素时丝裂原活化蛋白激酶的酪氨酸磷酸化水平增加了2.1±0.6倍。最后，制备了与生物素部分偶联的肽HC衍生物，结果显示其能与野生型胰岛素受体的β亚基以及羧基末端缺少43个氨基酸的截短受体结合。然而，该肽与IGF-1受体或表皮生长因子受体几乎没有结合（如果有结合的话）。综上所述，我们的数据表明与胰岛素受体羧基末端相关联的一个十五肽可与胰岛素受体β亚基结合，并且这种相互作用可能有助于增强受体的内在活性及信号转导。
