SULFHYDRYL AGENTS MODULATE INSULIN-RECEPTOR AND EPIDERMAL GROWTH-FACTOR (EGF)-RECEPTOR KINASE VIA REACTION WITH INTRACELLULAR RECEPTOR DOMAINS - DIFFERENTIAL-EFFECTS ON BASAL VERSUS ACTIVATED RECEPTORS
SULFHYDRYL AGENTS MODULATE INSULIN-RECEPTOR AND EPIDERMAL GROWTH-FACTOR (EGF)-RECEPTOR KINASE VIA REACTION WITH INTRACELLULAR RECEPTOR DOMAINS - DIFFERENTIAL-EFFECTS ON BASAL VERSUS ACTIVATED RECEPTORS
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DOI：
10.1042/bj2920217
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发表时间：
1993-05-15
影响因子：
4.1
通讯作者：
KONSTANTOPOULOS, N
中科院分区：
文献类型：
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作者：
CLARK, S;KONSTANTOPOULOS, N
巯基试剂已被证明对胰岛素受体的结构和功能产生各种影响。然而，还没有人尝试将这些效应定位于特定的受体结构域。我们已经用胰岛素和表皮生长因子(EGF)受体(两者都是受体酪氨酸激酶，但在外部结构域具有不同的硫基/二硫键结构)和胰岛素受体激酶(IRK)蛋白研究了这个问题，IRK蛋白仅由胰岛素受体胞浆结构域组成，并显示构成激酶活性。结果显示，基础受体和激活受体的反应不同。生理性还原剂GSH可刺激基础受体的自磷酸化，但不影响(EGF)或抑制(Ins)激活的受体，且受体结构未见明显减少。这些结果与二硫苏糖醇的结果相反，二硫苏糖醇似乎激活了与细胞外胰岛素受体二硫化物减少相关的磷酸化，但对EGF受体或irk蛋白没有影响。烷基化试剂N-乙基马来酰亚胺(NEM)和碘乙酰胺(IAM)对受体自磷酸化有相反作用。但只有在基础状态下，IAM才能保护受体免受NEM的抑制作用。我们的结果表明，在胰岛素和EGF受体的细胞质区域内可以发生复杂的巯基相互作用来改变受体激酶的活性。受体的基础状态和激活状态在硫代试剂作用方面是不同的，可能是由于配体(胰岛素、EGF)或结构性激活(IRK)导致受体构象发生变化。
Sulphydryl reagents have been shown to produce a variety of effects on insulin-receptor structure and function. However, localization of these effects to specific receptor domains has not been attempted. We have investigated this question with insulin- and epidermal growth factor (EGF)-receptors (both are receptor tyrosine kinases but have different sulphydryl/disulphide structures within the external domain), and the insulin receptor kinase (IRK) protein consisting solely of the insulin-receptor cytoplasmic domain and exhibiting constitutive kinase activity. Results showed a differential response between basal and activated receptors. The physiological reductant GSH stimulated basal receptor autophosphorylation, but was either without effect (EGF) or inhibited (insulin) activated receptors, and occurred without visible reduction of receptor structure. These results contrast with those obtained with dithiothreitol which appears to activate phosphorylation in association with reduction of the extracellular insulin-receptor disulphides, but is without effect on the EGF receptor or the IRK protein. Alkylating agents N-ethylmaleimide (NEM) and iodoacetamide (IAM) had opposing effects on receptor autophosphorylation. However, only in the basal state was IAM able to protect receptors from the inhibitory effect of NEM. Our results suggest that complex sulphydryl interactions can occur within the cytoplasmic domain of insulin- and EGF-receptors to alter receptor kinase activity. The basal and activated state of receptors is not the same with respect to sulphydryl reagent action, possibly due to conformational change in the receptor induced by ligand (insulin, EGF) or constitutive (IRK) activation.