NEUROTRANSMITTER RECEPTOR PURIFICATION AND STRUCTURE

神经递质受体；肾上腺素能(β1、β2、α1和 Alpha2)，胆碱能(毒碱和尼古丁)，以及 苯二氮卓类受体的分离纯化工作正在有序进行 为了了解受体功能的分子基础和 神经元通讯。具体项目正在进行中，以 提供上述每个项目的准确结构信息 受体蛋白。正在获得的结构数据包括主要数据 序列数据、蛋白水解图、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、膜结构域和效应器 偶联蛋白质识别结构域。我们的数据显示 证明了在结构上存在相似性的非 药理相关神经递质受体 (毒鼠碱、胆碱能和α肾上腺素能)和这些 神经递质受体通过 神经递质引发的蛋白质构象变化 结合到其跨膜结构域中的结合部位。 受体偶联是由细胞内环路介导的 受体，似乎是效应蛋白(GTP- 调节蛋白)受体的识别部分。电子 显微技术与高效尺寸排阻色谱 纯化的受体表明α2和β2受体 以同源二聚体形式存在，而M受体为单体。 已经建立了蛋白质制备程序，该程序 包括各种高效液相步骤、配基亲和层析 单抗亲和层析，制备性十二烷基硫酸钠- 凝胶电泳、凝集素无限层析、离子交换 净化方案现在允许同时详细地 所有肾上腺素能和胆碱能受体的结构比较 蛋白质以及从克隆的受体基因表达的蛋白质。 *(于1987年7月7日从LNP转来)。

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. 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 their transmembrane domains. Receptor coupling is mediated by the intracellular loops of the receptors which appears to be the effector protein (GTP- regulatory protein) recognition portion of the receptor. Electron microscopy and high 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 afinity chromatgraphy, ion exchange purification protocols is now permitting simultaneous detailed structural comparisons of all adrenergic and cholinergic receptor proteins as well as proteins expressed from cloned receptor genes. *(Transferred from LNP on 7/87).