STRUCTURE AND FUNCTION OF NICOTINIC RECEPTORS

描述(改编自应用)：本研究的长期目标 建议将电生理、生化、生物物理和 深入了解其结构和功能的分子生物学技术 烟碱型乙酰胆碱受体(AChR)的研究进展 蛋白质-脂质相互作用。这个项目的重点是组织在三个方面 独立方法：(1)第一种方法是扩展正在进行的站点 对鱼雷AChR的脂质暴露残基进行定向突变 显示为影响通道门控机构。调查人员提议 在新的疏水变构上引入额外的氨基酸取代 定位阿尔法-V425定位。这些研究将进一步检验这一假设 脂蛋白界面上的一些变构位点具有 在AChR通道门控中的关键作用。身份识别与功能 对这些脂类暴露的变构部位的表征将提供进一步的 对AChR的结构-功能关系的洞察。(2)第二个 一种方法建议引入周期性色氨酸替代，以覆盖所有 沿M4跨膜区的β、γ和增量的位置 亚单位。对一系列色氨酸取代的系统分析将 提供螺旋-螺旋或亚基-亚基的物理位置信息 对于AChR的功能性齐聚至关重要的接触。这个 可能的约束位置的位置可以识别附加的 关于M4跨膜结构域的空间组织的方面 对M1和M3结构域以及结构-功能关系的影响 Achr.(3)第三种方法是利用新的鱼雷M4 AChR突变 检查脂质-蛋白质界面对各种不同的作用的敏感性 全身麻醉药的种类。本项目的目标是定义 这些变构位点的生化和生物物理相互作用 确定M4跨膜的空间组织的其他方面 AChR的结构域。

Description (Adapted from Application): The long-range goal of this research proposal is to combine electrophysiological, biochemical, biophysical and molecular biological techniques to gain insight into the structure and function of the nicotinic acetylcholine receptor (AChR) with special attention on protein-lipid interactions. The emphasis of this project is organized on three independent approaches: (1) The first approach is to extend ongoing site directed mutagenesis at lipid-exposed residues of the Torpedo AChR that have shown to affect the channel gating mechanism. The investigators propose to introduce additional amino acid replacements at the new hydrophobic allosteric position alpha-V425 position. These studies will further test the hypothesis that a number of allosteric sites at the lipid protein interface have a critical role on the AChR channel gating. The identification and functional characterization of these lipid-exposed allosteric sites will provide further insights into the structure-function relationships of the AChR. (2) The second approach proposes to introduce periodic tryptophan substitutions to cover all the positions along the M4 transmembrane domains for beta, gamma and delta subunits. A systematic analysis of a series of tryptophan substitutions will provide information of the physical location of helix-helix or subunit-subunit contacts that are critical for the functional oligomerization of the AChR. The positional location of possible constraint sites may identify additional aspects of the spatial organization of the M4 transmembrane domains in respect to the Ml and M3 domains as well as to structure-function relationships of the AChR. (3) The third approach is to use novel Torpedo M4 AChR mutations to examine the sensitivity of the lipid-protein interface to the action of various types of general anesthetics. The objectives of this project are to define the biochemical and biophysical interactions of these allosteric sites and to identify additional aspects of the spatial organization of the M4 transmembrane domains of the AChR.