HORMONE REGULATED GTP-BINDING PROTEINS

激素调节的 GTP 结合蛋白

• 批准号: 2608407
• 负责人: JOHN D HILDEBRANDT
• 金额: $ 25.11万
• 依托单位: MEDICAL UNIVERSITY OF SOUTH CAROLINA
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-07-01 至 1999-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2608407
• 关键词: G protein; SDS polyacrylamide gel electrophoresis; active sites; biological polymorphism; biological signal transduction; chemical association; dimer; guanine nucleotide binding protein; high performance liquid chromatography; hormone receptor; hormone regulation /control mechanism; immunoprecipitation; laboratory rabbit; membrane proteins; molecular cloning; posttranslational modifications; protein biosynthesis; protein isoforms; protein purification; protein sequence; protein structure; protein structure function; receptor coupling; recombinant proteins; site directed mutagenesis

Reversible G protein subunit interactions will be studied to determine the properties of these proteins important for mediating hormone action. G proteins are heterotrimeric GTP binding proteins composed of alpha, beta and gamma subunits. They mediate signalling across the plasma membranes of cells for many hormones and neurotransmitters, as well as for numerous other regulatory substances. External signals activate G proteins inside cells by stimulating GTP-dependent dissociation of their alpha subunit from their associated betagamma dimer. These parts of the G protein then independently control signalling pathways inside cells so that the cells respond appropriately to signals in their environment. Errors in such signalling systems are likely contributors to disease processes such as diabetes, essential hypertension and cancer. Based upon the fact that some hormones require very specific G protein isoform combinations for their effects, it is hypothesized that G protein subunit dissociation and reassociation during hormone signalling provides a key mechanism for coordinating cell responses. For example, if G protein subunits form transient or dynamic heterotrimeric complexes, hormones will only be able to stimulate cells when G proteins containing the correct subunits are present. To test this hypothesis the regions of betagamma dimers that bind G protein alpha subunits will be identified and this information will be used to develop sensitive and general assays for G protein activation by receptors in intact membranes and cells. The Specific Aims of this application are: 1). To determine the structure and diversity of naturally occurring gamma subunits in G protein heterotrimers. (2). To determine the post-translational modifications of the beta subunits of the G proteins. 3). To use the information from 1) and 2) to define regions in G protein subunits involved in the binding of betagamma dimers to alpha subunits. 4). To determine if G protein heterotrimers are formed of specific subunit combinations in cells and whether reversible subunit dissociation produces heterotrimers of changing subunit composition. 5). To determine the occurrence, regulation and function of G protein subunit dissociation in intact membranes using fluorescently-labeled peptides based upon the work in Specific Aims 1-3. These studies will test the hypothesis that changing G protein subunit composition alters hormonal responses of cells. Regardless of the validity of this hypothesis, these studies will define the properties of the G proteins important for the action of the hormones that regulate them.

将研究可逆的G蛋白亚基相互作用，以确定 这些蛋白质的性质对介导激素作用很重要。 G 蛋白质是由α，β 和γ亚基。 它们介导跨质膜的信号传导 细胞的许多激素和神经递质，以及许多 其他管制物质。 外部信号激活内部的G蛋白 通过刺激α亚单位的GTP依赖性解离 从它们的β-γ二聚体中分离出来。 G蛋白的这些部分 独立控制细胞内的信号通路， 对环境中的信号做出适当的反应。 错误在这样的 信号系统可能是疾病过程的贡献者， 糖尿病、原发性高血压和癌症。 基于以下事实 某些激素需要非常特异的G蛋白同种型组合， 它们的作用，假设G蛋白亚基解离和 激素信号传导过程中的再结合提供了 协调细胞反应。 例如，如果G蛋白亚基形成 瞬时或动态异源三聚体复合物，激素将只能 当含有正确亚基的G蛋白被 礼物 为了验证这一假设，我们研究了β-γ二聚体的 结合G蛋白α亚基将被识别，这些信息将 用于开发G蛋白激活的敏感和通用测定 通过完整的细胞膜和细胞中的受体。 具体目标是 应用：1）。确定的结构和多样性 G蛋白异源三聚体中天然存在的γ亚基。 （二）、到 确定蛋白质的β亚基的翻译后修饰， G蛋白。 3）。使用1）和2）中的信息来定义区域 在参与β-γ二聚体与α-γ二聚体结合的G蛋白亚基中， 亚单位。 4）。为了确定G蛋白异源三聚体是否由 细胞中的特定亚基组合以及是否可逆亚基 解离产生改变亚基组成的异源三聚体。 5）。 确定G蛋白亚基的发生、调节和功能 使用荧光标记肽在完整膜中解离 根据具体目标1-3中的工作。 这些研究将测试 改变G蛋白亚基组成改变激素水平的假说 细胞的反应。 不管这个假设的正确性如何， 研究将确定G蛋白的重要特性， 调节它们的激素的作用。