HORMONE REGULATED GTP-BINDING PROTEINS

一类鸟嘌呤核苷酸相互作用的机制 作为中介的结合蛋白(G蛋白) 细胞表面激素受体及其细胞内效应因子 系统将被调查。至少有四个不同的成员 在这个家族中，大多数有多个同工酶，用来协调 至少六种不同的细胞内调节因子的活性 小路。这些蛋白质调节重要的激素或激素样蛋白。 基本上对所有哺乳动物细胞都有影响。他们都有一个相似的 亚基组成并共享一个公共亚单位。此应用程序 建议调查这个公共亚单位是如何可逆地关联的 与蛋白质的其他亚基结合，并通过 它们相互影响的功能，从而协调 细胞内调节通路。这有五个组成部分 项目。(特定目标1)完整亚基相互作用的研究 培养中生长的s49小鼠淋巴瘤细胞膜。这些研究 将表明亚基解离在 未用洗涤剂从膜上增溶的蛋白质 并随后从其他可能调节蛋白质的蛋白质中纯化出来 它们的功能。(特定目标2)G亚基解离的研究 从牛脑或人红细胞中提纯的蛋白质或类似物 重组蛋白，在洗涤剂溶液中；这将决定什么 调节不同蛋白质之间的相互作用 亚单位。(具体目标3)研究生物多样性的功能后果 纯化的G蛋白的可逆亚单位相互作用 通过检测GTP结合动力学确定的洗涤剂溶液 和对纯化的蛋白质的水解性。(具体目标4)研究 这些可逆亚单位相互作用如何影响特定的 S49细胞完整膜的细胞内调节系统， CAMP-腺苷环化酶系统。(具体目标5)确定 蛋白质上的亚基结合部位，主要由 分离与蛋白质化学修饰有关的片段 阻止亚基相互作用。这些研究将提供重要的 关于这些蛋白质的结构信息及其亚基如何 互动。

The mechanisms for the interactions of a family of guanine nucleotide binding proteins (G proteins) which act as intermediaries between hormone receptors on the cell surface and their intracellular effector systems will be investigated. There are at least four distinct members of this family, most with multiple isoforms, that serve to coordinate the activities of at least six different intracellular regulatory pathways. These proteins mediate important hormone or hormone-like effects in essentially all mammalian cells. They all have a similar subunit composition and share one common subunit. This application proposes to investigate how this common subunit reversibly associates with the other subunits of the proteins and provides a mechanism by which they affect each others function, thereby coordinating intracellular regulatory pathways. There are five components of this project. (Specific Aim 1) The study of subunit interactions in intact membranes from s49 mouse lymphoma cells grown in culture. These studies will indicate the role of subunit dissociation in the behavior of proteins which have not been solubilized from membranes with detergents and subsequently purified away from other proteins which may modulate their function. (Specific Aim 2) The study of subunit dissociation of G proteins purified from bovine brain or human erythrocytes, or analogous recombinant proteins, in detergent solution; which will determine what regulates the interactions of the different proteins with their shared subunit. (Specific Aim 3) The study of the functional consequences of these reversible subunit interactions of purified G proteins in detergent solution, determined by examining the kinetics of GTP binding and hydrolysis to the purified proteins. (Specific Aim 4) The study of how these reversible subunit interactions affect a specific intracellular regulatory system in intact membranes from s49 cells, the CAMP-adenylyl cyclase system. (Specific Aim 5) The identification of the subunit binding sites on the proteins, determined primarily by isolating chemically-modified fragments of the proteins responsible for blocking subunit interactions. These studies will provide important structural information about these proteins and how their subunits interact.