BINDING SITES OF THE ACETYLCHOLINE ACCEPTOR

尼古丁乙酰胆碱受体含有配体门控离子通道 介导神经肌肉的化学-电传导 结合从神经末梢释放的乙酰胆碱。的 这项建议的广泛的长期目标是了解如何 激动剂结合诱导通道开放和构象变化， 受体。具体来说，电荷-电荷相互作用在 乙酰胆碱受体功能将得到系统研究。的 具体目标是：确定静电相互作用受体 将系统研究其功能。具体目标是： 确定激动剂之间的静电相互作用。具体目标 是：确定激动剂与其 结合位点，以及带电基团是否存在于该位点以及如何存在 它们分布;以确定静电相互作用对 乙酰胆碱受体的构象平衡，以及 静电相互作用负责构象转换; 并确定静电相互作用的作用， 竞争性阻断剂（NCB）和通道位点，以及 不同构象的离子通道分布不同。的 激动剂和NCB的平衡、缔合和解离常数 将通过常规的荧光结合测量来确定， 不同离子强度下的停流荧光光谱。这些 测量也将表征当地的人口 构象和构象转变速率。乙酰胆碱 受体是疾病肌无力自身免疫抗体的目标 重症和局部麻醉药。拟议的研究将有助于确定 这些结构域相对于配体结合的结构，因此将 有助于合理的药物设计。

The nicotine acetylcholine receptor contains a ligand-gated ion channel which mediates the chemo-electrical transduction at the neuromuscular junction by binding acetylcholine released from the nerve terminal. The broad long-term objectives of this proposal is to understand how the agonist-binding induces the channel opening and conformational changes of the receptor. Specifically, the role of charge-charge interactions in acetylcholine receptor function will be systematically studied. The specific aims are: to determine electrostatic interactions receptor function will be systematically studied. The specific aims are: to determine electrostatic interactions between agonist. The specific aims are: to determine electrostatic interactions between agonist and its binding sites, and whether the charged groups exist at the sites and how they distribute; to determine the effect of electrostatic interactions on the conformation equilibrium of the acetylcholine receptor, and whether electrostatic interactions are responsible for conformational conversions; and to determine the role of electrostatic interactions between non- competitive blocker (NCB) and the channel site, and whether the changes in the ion channel distribute differently for different conformations. The equilibrium, association and dissociation constants of agonist and NCB will be determined by conventional fluorescence binding measurements and stopped flow fluorescence spectroscopy at various ionic strength. These measurements will also characterize the populations of the native conformations and rate of conformational transitions. The acetylcholine receptor is the target of auto-immune antibodies in the disease myasthenia gravis and local anesthetics. The proposed research will help define the structure of those domains relative to ligand binding and thus will contribute to rational drug design.