Structure-function of the α4β2 nicotinic acetylcholine receptor in a lipidic environment

项目摘要(1-2段)： 烟碱型乙酰胆碱受体是配体门控离子通道，广泛表达于 中枢和外周神经系统。α-4-β-2烟碱受体是大脑中含量最丰富的亚型 它与尼古丁成瘾和包括阿尔茨海默氏症和帕金森氏症在内的神经疾病有关。 尼古丁慢性暴露可上调α-4-β-2受体在体外和小鼠脑内的表达 吸烟者与不吸烟者的比较。因此，已经作出了广泛的努力来发展小型 通过靶向这种受体类型来减少烟草成瘾的分子，但这些努力受到 尼古丁受体的结构信息量有限。一种钝化态烟碱的原子结构 通过X射线结晶学和低温电子显微镜获得的洗涤剂中的受体是已知的，但高分辨率结构 在生理相关的脂类环境中仍未解决。此外，我们缺乏结构性信息， 这种受体在门控周期中处于不同的状态。在这里，我建议首先确定脂类的组合 使用电生理方法支持健壮的通道功能。同时，我将优化样品 利用功能信息在脂类纳米盘中制备α4β2烟碱受体 在特定的脂类环境中进行研究。我将利用这些准备工作来获得高分辨率的结构 α4β2烟碱受体在不同构象状态的信息，在实验条件下 直接进行生理准备。膜上和不同细胞中受体的结构信息 构象状态将为理解这些受体的变构门控提供试金石 用于开发国家选择性药理工具和治疗学的模板。

Project summary (1-2 para): Nicotinic acetylcholine receptors are ligand-gated ion channels that are widely expressed throughout the central and peripheral nervous systems. The α4β2 nicotinic receptor is the most abundant subtype in the brain and is linked to nicotine addiction and neurological disorders including Alzheimer's and Parkinson's diseases. Expression of the α4β2 receptor is upregulated by chronic exposure to nicotine in vitro and in the brains of smokers compared to that of non-smokers. Accordingly, there have been extensive efforts to develop small molecules to reduce tobacco addiction by targeting this receptor type, but these efforts are hampered by the limited amount of structural information for nicotinic receptors. Atomic structures of a desensitized-state nicotinic receptor in detergent obtained by X-ray crystallography and Cryo-EM are known, but high-resolution structures in a physiologically relevant lipidic environment remain unresolved. Moreover, we lack structural information for this receptor in the different states along the gating cycle. Here I propose to first identify combinations of lipids that support robust channel function using electrophysiological approaches. In parallel I will optimize sample preparation for the α4β2 nicotinic receptor in lipidic nanodiscs, leveraging the information from the functional studies in defined lipidic environments. I will use these preparations to obtain high resolution structural information for the α4β2 nicotinic receptor in distinct conformational states, in an experimental condition tied directly to a physiological preparation. Structural information for the receptor in a membrane and in different conformational states will provide touchstones for understanding allosteric gating of these receptors and templates for development of state-selective pharmacological tools and therapeutics.