Binding of G protein beta gamma-subunits to pleckstrin homology domains.
Binding of G protein beta gamma-subunits to pleckstrin homology domains.
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发表时间：
1994-04
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通讯作者：
K. Touhara;James Inglese;J. Pitcher;G. Shaw;R. Lefkowitz
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作者：
K. Touhara;James Inglese;J. Pitcher;G. Shaw;R. Lefkowitz
许多受体的配体诱导的活化导致异源三聚体G蛋白的α-和β γ-亚基复合物的解离，这两者都调节参与细胞信号传导过程的各种效应分子。在一种情况下，胞质酶β-肾上腺素能受体激酶（β方舟）结合异源三聚体G蛋白（G β γ）的解离的、异戊二烯化的、膜锚定的β γ亚基，从而靶向其膜结合受体底物。最近，许多参与细胞信号转导的蛋白质已经显示含有与最初在蛋白质“普列克底物蛋白”中鉴定的“结构域”（普列克底物蛋白同源结构域; PH结构域）同源的序列，并且随后在β方舟序列的G β γ相互作用区域中发现。在这里，我们证明了谷胱甘肽S-转移酶融合蛋白（包含该组九种蛋白质的PH结构域的序列）在不同程度上结合G β γ。通过直接结合试验或通过阻断G β γ介导的β ARK 1膜易位的能力，记录G β γ与这些融合蛋白的结合。G β γ与这些融合蛋白的结合被Go的α亚基（Go α）抑制，表明G β γ与G α和含有PH结构域的融合蛋白的结合是相互排斥的。对一系列来自Ras-鸟嘌呤-核苷酸释放因子的截短PH结构域的研究表明，G β-γ结合结构域仅包括PH结构域的C-末端部分和刚好在其远端的序列。G β γ和PH结构域蛋白质之间的蛋白质-蛋白质相互作用可能在细胞信号传导中起重要作用，类似于先前对Src同源性2和3结构域所证明的作用。
Ligand-induced activation of many receptors leads to dissociation of the alpha- and beta gamma-subunit complexes of heterotrimeric G proteins, both of which regulate a variety of effector molecules involved in cellular signaling processes. In one case, a cytosolic enzyme, the beta-adrenergic receptor kinase (beta ARK) binds to the dissociated, prenylated, membrane-anchored beta gamma-subunits of heterotrimeric G proteins (G beta gamma) and is thereby targeted to its membrane-bound receptor substrate. Quite recently, numerous proteins involved in cellular signal transduction have been shown to contain sequences homologous with a "domain" originally identified in the protein "pleckstrin" (pleckstrin homology domain; PH domain) and subsequently found in the G beta gamma interaction region of the beta ARK sequence. Here we demonstrate that glutathione S-transferase-fusion proteins, containing sequences encompassing the PH domain of nine proteins from this group, bind G beta gamma to varying extents. Binding of G beta gamma to these fusion proteins was documented either by a direct binding assay or by ability to block G beta gamma-mediated membrane translocation of beta ARK1. G beta gamma binding to these fusion proteins was inhibited by the alpha subunit of Go (Go alpha), indicating that the binding of G beta gamma to G alpha and the PH domain-containing fusion proteins is mutually exclusive. Studies with a series of truncated PH domains derived from the Ras-guanine-nucleotide-releasing factor indicate that the G beta gamma binding domain includes only the C-terminal portion of the PH domain and sequences just distal to this. Protein-protein interactions between G beta gamma and PH domain-containing proteins may play a significant role in cellular signaling analogous to that previously demonstrated for Src homology 2 and 3 domains.