3D Structure and Function of Neuronal Nicotinic Acetylcholine Receptors

项目摘要：神经型烟碱型乙酰胆碱受体(NAChRs)是五聚体配基门控离子 介导快速突触传递的通道，是糖尿病的重要和新的治疗靶点 神经退行性疾病和精神疾病。目前还没有原子分辨率结构用于任何 烟碱受体，以及这个Cys-loop受体超家族中的NO受体，其结构特征是 不止一种构象。这项研究的长期目标是更好地了解 构象变化是不同功能状态之间转换的基础，为了发展可靠的， 与结构导向药物设计相关的nAChRs的原子分辨模型。为了实现这些目标， 将采用以下方法。1.通过受体同源基因筛选，筛选出7个候选受体 结晶，以及优化洗涤剂中受体稳定性的测试条件。2.使用快速灌流补片 钳制和单通道分析，测试结构修饰和药理探针的组合 稳定不同的受体构象。3.结晶不同构象状态的受体和 测定(A)封闭休眠受体、(B)开放激活受体和(C)a的原子分辨结构 闭合脱敏受体。结构研究将得到功能分析的补充，测试新的 关于受体中状态转换机制的基于结构的假设。结果将会有 对一般的Cys-loop受体家族和特定的7个nAChRs的广泛影响。

Project Summary: Neuronal nicotinic acetylcholine receptors (nAChRs) are pentameric ligand-gated ion channels that mediate fast synaptic transmission and are important and novel therapeutic targets in neurodegenerative diseases and mental illness. Currently there is no atomic-resolution structure for any nicotinic receptor, and no receptor in this Cys-loop receptor superfamily has been structurally characterized in more than one conformation. The long-term objectives of the research are to better understand the conformational changes that underlie the transitions between different functional states and to develop reliable, atomic-resolution models of nAChRs relevant to structure-guided drug design. Toward these goals the following approach will be used. 1. Through receptor ortholog screening, identify an ¿7 receptor candidate for crystallization, and test conditions to optimize receptor stability in detergent. 2. Using fast-perfusion patch clamp and single channel analysis, test combinations of construct modifications and pharmacological probes to stabilize distinct receptor conformations. 3. Crystallize receptor in different conformational states and determine atomic-resolution structures of (a) a closed-resting receptor, (b) an open-activated receptor and (c) a closed-desensitized receptor. The structural studies will be complemented with functional assays that test new structure-based hypotheses regarding mechanisms of state transitions in the receptor. The results will have broad implications for the Cys-loop receptor family in general and ¿7 nAChRs specifically.