Epidermal growth factor (urogastrone)-mediated phosphorylation of a 35-kDa substrate in human placental membranes: relationship to the beta subunit of the guanine nucleotide regulatory complex.
Epidermal growth factor (urogastrone)-mediated phosphorylation of a 35-kDa substrate in human placental membranes: relationship to the beta subunit of the guanine nucleotide regulatory complex.
复制标题
表皮生长因子（尿胃素）介导的人胎盘膜中 35 kDa 底物的磷酸化：与鸟嘌呤核苷酸调节复合物的 β 亚基的关系。
DOI：
10.1073/pnas.83.2.236
复制
发表时间：
1986
影响因子：
11.1
通讯作者：
M. Hollenberg
中科院分区：
文献类型：
--
作者：
K. A. Valentine;J. Northup;M. Hollenberg
我们已经鉴定出人胎盘膜制剂中存在约 35 kDa (pp35) 的成分，它是表皮生长因子（尿胃素）[EGF(Uro)] 介导的磷酸化的底物。 EGF(Uro) 刺激的 pp35 磷酸化是钙依赖性的，并且在钙存在（但不存在）的情况下制备的膜中显着增强。在 EGF(Uro) 存在下掺入 pp35 的磷酸盐是碱稳定的，并且以 O4-磷酸酪氨酸的形式存在。在相同条件下，胰岛素不刺激 pp35 磷酸化。无论是天然形式还是磷酸化形式，pp35 在钙螯合剂 (EDTA/EGTA) 存在下都可以从膜上释放；并通过在钙存在下将 EDTA/EGTA 洗出液加回 EDTA/EGTA 洗涤的膜来重建 EGF(Uro) 刺激的磷酸化。 pp35 的特性与 beta-35 的特性即使不完全相同，也是相似的，β-35 是一种 35 kDa 的多肽，与刺激 (Gs) 或抑制 (Gi) 腺苷酸环化酶系统的鸟嘌呤核苷酸结合寡聚物的 β 亚基相似。与 pp35 一样，在使用 EDTA/EGTA 洗涤的胎盘膜或固定在伴刀豆球蛋白 A-琼脂糖上的溶解的 EGF(Uro) 受体的重建系统中观察到 EGF(Uro) 刺激的离体兔肝 beta-35 的磷酸化。相反，添加源自兔肝 Gi 或牛转导蛋白的 β 亚基不会导致重构系统中 35-kDa 底物的磷酸化。此外，胎盘膜释放的 35 kDa 蛋白质与抗转导蛋白抗体发生交叉反应，该抗体可以识别从多种来源分离的 β 亚基。我们得出的结论是，人胎盘 pp35 底物可能代表 β-35 蛋白的胎盘等价物。我们的数据表明参与生长因子作用的受体与利用 GTP 结合蛋白作为转导元件的调节系统之间可能存在联系。
We have identified a component of about 35 kDa (pp35), present in human placental membrane preparations, that is a substrate for epidermal growth factor (urogastrone) [EGF(Uro)]-mediated phosphorylation. The EGF(Uro)-stimulated phosphorylation of pp35 was calcium-dependent and was markedly enhanced in membranes prepared in the presence (but not in the absence) of calcium. The phosphate incorporated into pp35 in the presence of EGF(Uro) was alkali-stable and was present as O4-phosphotyrosine. Under identical conditions, insulin did not stimulate pp35 phosphorylation. Either in its native or in its phosphorylated form, pp35 could be released from the membranes in the presence of calcium-chelating agents (EDTA/EGTA); and EGF(Uro)-stimulated phosphorylation was reconstituted by adding back EDTA/EGTA eluates to EDTA/EGTA-washed membranes in the presence of calcium. The properties of pp35 were similar if not identical to those of beta-35, a 35-kDa polypeptide similar to the beta subunit of the guanine nucleotide-binding oligomers that stimulate (Gs) or inhibit (Gi) the adenylate cyclase system. As with pp35, EGF(Uro)-stimulated phosphorylation of isolated rabbit liver beta-35 was observed in a reconstituted system using either EDTA/EGTA-washed placental membranes or solubilized EGF(Uro) receptor immobilized on concanavalin A-agarose. In contrast, the addition of beta subunits derived from rabbit liver Gi or bovine transducin did not result in phosphorylation of a 35-kDa substrate in the reconstituted system. Further, a 35-kDa protein released from placental membranes crossreacted with an anti-transducin antibody that can recognize the beta subunit isolated from a variety of sources. We conclude that the human placental pp35 substrate likely represents the placental equivalent of the beta-35 protein. Our data point to a possible link between those receptors involved in growth-factor action and the regulatory systems that utilize GTP-binding proteins as transducing elements.