MUSCARINIC RECEPTOR COUPLING MECHANISMS

毒蕈碱受体偶联机制

• 批准号: 3405932
• 负责人: GARY R LUTHIN
• 金额: $ 9.01万
• 依托单位: HAHNEMANN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-07-01 至 1992-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3405932
• 关键词: adenylate cyclase; biological signal transduction; brain mapping; enzyme mechanism; guanine nucleotide binding protein; inhibitor /antagonist; laboratory rabbit; laboratory rat; muscarinic receptor; neuropharmacology; receptor coupling; receptor expression; stimulant /agonist

The long-term objectives of this project are to study mechanisms of signal transduction in the brain. A first step to understanding this process is t develop biochemical methodologies allowing molecular characterization of receptor and effector entities. To this end, antibodies to neurotransmitte receptors and guanine nucleotide binding proteins (G proteins) have been developed, taking advantage of recent elucidation of their primary amino ac d sequences. The antibodies,are used in an immunoaffinity purification of biologically active complexes of muscarinic receptor and G protein. Specifically, muscarinic receptors are labelled in cellular membranes with agonists or antagonists, then solubilized from the membranes. The receptor are purified over a lectin affinity resin, then immunoprecipitated with anti-receptor antibodies. In the presence of agonist, this allows co-purification of associated G proteins. The G proteins present in comple with the receptor are identified using monospecific antibodies to their subunits. This experimental design will be used to identify muscarinic receptor-G protein complexes in various brain regions, and is of interest because of the recent cloning of at least 5 subtypes of the receptor, most of which can be found in any of several brain regions. The experiments can resolve the issue of which G protein each receptor subtype interacts with i a given brain region. The combination of different receptor subtypes, and a potential of each subtype to interact with any of several G proteins, cou d be the molecular basis for the apparent specificity of cellular responses t receptor agonists. A dysfunction of cellular signal transduction could occ r as a result of inappropriate expression of a receptor or G protein subtype, with a consequent subordination off normal signal pathways. This could in turn form the basis for disease states such as Altzheimer's, and for learni g and memory deficits where the cholinergic pathways in brain are believed to be altered.

该项目的长期目标是研究信号的机制 大脑中的信号传导。 理解这个过程的第一步是 开发生物化学方法，允许分子表征 受体和效应子实体。 为此，神经递质抗体 受体和鸟嘌呤核苷酸结合蛋白（G蛋白）已经被 开发，利用最近阐明其伯氨基酸 D 序列的 抗体用于免疫亲和纯化 毒蕈碱受体和G蛋白的生物活性复合物。 具体地，毒蕈碱受体在细胞膜中被标记， 激动剂或拮抗剂，然后从膜上溶解。 受体 在凝集素亲和树脂上纯化，然后用 抗受体抗体 在激动剂的存在下，这允许 相关G蛋白的共纯化。 G蛋白存在于复杂的 使用针对其受体的单特异性抗体来鉴定其 亚单位。 本实验设计将用于鉴定毒蕈碱 受体-G蛋白复合物在不同的大脑区域，并感兴趣的是 由于最近克隆了该受体的至少5种亚型， 在大脑的几个区域都能找到。 这些实验可以 解决每个受体亚型与哪种G蛋白相互作用的问题 一个特定的大脑区域。 不同受体亚型的组合，以及 每种亚型与几种G蛋白中的任何一种相互作用的潜力， D 是细胞反应表观特异性的分子基础， 受体激动剂 细胞信号转导功能障碍可能会发生。 R 由于受体或G蛋白亚型的不适当表达， 从而导致正常信号通路的从属性。 这可以在 转形的基础上，疾病的国家，如阿尔图，并为学习 G 和记忆缺陷，其中大脑中的胆碱能途径被认为是 被改变。