NEUROTRANSMITTER RECEPTOR PURIFICATION AND STRUCTURE

神经递质受体的纯化和结构

• 批准号: 3969050
• 负责人: J C VENTER
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF NEUROLOGICAL DISORDERS AND STROKE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3969050
• 关键词: Alzheimer's disease; Huntington's disease; affinity chromatography; benzodiazepines; conformation; electrofocusing; electron microscopy; electrophysiology; gel electrophoresis; guanosine triphosphate; high performance liquid chromatography; human tissue; immunochemistry; ion exchange chromatography; liposomes; monoclonal antibody; neural conduction; proteolysis

Neurotransmitter receptors; adrenergic (beta1, beta2, alpha1 and alpha2), cholinergic (muscarinic and nicotinic), and benzodiazepine receptors are being isolated and purified in order to understand the molecular basis of receptor function and neuronal communication. Specific projects are underway to provide precise structural information on each of the above receptor proteins. Structural data being obtained include primary sequence data, proteolytic digest maps, topology information and structure-function data, e.g., neurotransmitter binding site localization, sugar localization, membrane domain and effector coupling protein recognition domains. The structure of receptors from Alzheimer's and Huntington's disease patients are being examined by immunological techniques. Our data have demonstrated that structural similarities exist among non-pharmacologically related neurotransmitter receptors (muscarinic, cholinergic and alpha adrenergic) and that these neurotransmitter receptors mediate cellular modulation via protein conformational changes initiated by neurotransmitter binding to the binding site in the extracellular protein domain. Receptor coupling is mediated by the cytoplasmic "tail" of the receptors which appears to be the effector protein (GTP-regulatory protein) recognition portion of the receptor. Electron microscopy, performance size-exclusion chromatography of purified receptors indicate that the alpha2 and beta2 receptors exist as homodimers while the muscarinic receptor is monomeric. Protein preparative procedures have been established which include various HPLC steps, ligand affinity chromatography, monoclonal antibody affinity chromatography, preparative SDS-gel electrophoresis, lectin affinity chromatography, ion exchange chromatography and column isoelectric focusing. The establishment of these purification protocols is now permitting simultaneous detailed structural comparisons of all adrenergic and cholinergic receptor proteins.

神经递质受体;肾上腺素能（β 1、β 2、α 1和α 2）， 胆碱能（毒蕈碱和烟碱）和苯二氮卓受体是 分离和纯化，以了解分子基础， 受体功能和神经元通讯。 具体项目 正在进行中，以提供上述各项的精确结构信息 受体蛋白 获得的结构数据包括一级层序 数据、蛋白水解消化图谱、拓扑信息和结构-功能 数据，例如，神经递质结合位点定位，糖定位， 膜结构域和效应偶联蛋白识别结构域。 的 阿尔茨海默病和亨廷顿病患者受体的结构 正在接受免疫学技术的检查 我们的数据表明， 非毒蕈碱相关的神经递质受体（毒蕈碱， 胆碱能和α肾上腺素能），这些神经递质受体 通过蛋白质构象变化介导细胞调节， 神经递质与细胞外蛋白结合位点的结合 域 受体偶联是由细胞质的“尾”介导的。 似乎是效应蛋白（GTP调节蛋白）的受体 受体的识别部分。 电子显微镜，性能 纯化受体的尺寸排阻色谱表明， α 2和β 2受体以同源二聚体存在，而毒蕈碱受体 受体是单体。 已经建立了蛋白质制备程序，其包括各种 HPLC步骤，配体亲和色谱，单克隆抗体亲和 层析，制备型SDS-凝胶电泳，凝集素亲和 色谱法、离子交换色谱法和柱等电 专注 这些纯化方案的建立现在是 允许同时对所有肾上腺素能受体进行详细的结构比较 和胆碱能受体蛋白。