EXPRESSION AND MUTAGENESIS OF CLONED ALPHA-2 RECEPTORS

克隆的 ALPHA-2 受体的表达和诱变

• 批准号: 2221122
• 负责人: LEE E LIMBIRD
• 金额: $ 21.04万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-01-01 至 1994-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2221122
• 关键词: adenylate cyclase; adrenergic receptor; computer assisted sequence analysis; enzyme inhibitors; eukaryote; gene deletion mutation; guanosine triphosphate; hydrogen channel; immunoprecipitation; ion exchange chromatography; ion transport; molecular cloning; molecular genetics; nucleic acid sequence; protein structure function; receptor binding; receptor expression; site directed mutagenesis; sodium channel; swine; transfection

The present studies are aimed at examining, using a molecular biological approach, the structural basis for the diverse functional properties of alpha2-adrenergic receptors. We plan to clone the porcine gene that codes for the alpha2-adrenergic receptor that we have purified to homogeneity. We will exploit insights gained from domain mapping and biochemical analysis of the purified receptor to delineate those areas of the receptor that we can analyze further by deletion and site-directed mutagenesis to learn what structural components are involved in ligand recognition, allosteric effects on adrenergic binding by Na+, H+ and 5-amino-substituted analogs of amiloride, receptor-GTP binding protein interactions, receptor-accelerated Na+/H+ exchange and receptor mediated inhibition of adenylate cyclase. Furthermore, we plan to examine what role different functional domains of the receptor play in the overall physiological functions influenced by alpha2-adrenergic receptors, such as suppression of neurotransmitter release, by expressing mutated versus wild type receptors in appropriate target cells to determine what effect discrete deletion or site mutations have on particular alpha2-receptor functions. We anticipate that the proposed studies will provide new insights into the structural basis for many of the functional properties of alpha2-receptors and perhaps suggest novel loci for drug development in mimicking, or blocking, the effects of alpha2-adrenergic agents in particular physiological processes.

目前的研究旨在使用分子生物学方法来检查 方法，不同功能特性的结构基础 α2-肾上腺素能受体。 我们计划克隆编码的猪基因 对于我们已经纯化至均质的α2-肾上腺素受体。 我们将利用从域映射和生化中获得的见解 分析纯化的受体以描绘受体的这些区域 我们可以通过删除和定点突变进一步分析 了解哪些结构成分参与配体识别， Na+、H+ 和 5-氨基取代对肾上腺素能结合的变构效应 阿米洛利类似物、受体-GTP 结合蛋白相互作用、 受体加速 Na+/H+ 交换和受体介导的抑制 腺苷酸环化酶。 此外，我们计划研究不同的角色 受体的功能域在整个生理学中发挥作用 受α2-肾上腺素能受体影响的功能，例如抑制 通过表达突变型与野生型受体来释放神经递质 在适当的靶细胞中确定离散缺失或 位点突变对特定的α2受体功能具有影响。 我们预计 拟议的研究将为结构性问题提供新的见解 α2 受体许多功能特性的基础，也许 提出了模拟或阻断药物开发的新位点 α2-肾上腺素能药物在特定生理过程中的作用。