Mapping the alcohol site of a neuronal potassium channel

绘制神经元钾通道的酒精位点

基本信息

批准号：
7760976
负责人：
MANUEL L COVARRUBIAS
金额：
$ 24.17万
依托单位：
THOMAS JEFFERSON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1997
资助国家：
美国
起止时间：
1997-01-01 至 2012-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7760976
关键词：
Acute Address Alanine Alcohol dependence Alcoholic Intoxication Alcohols Amino Acid Substitution Anesthetics Architecture Attention Binding Binding Sites Brain C-terminal Chemosensitization Coupling Development Drosophila genus Electrophysiology (science)Engineering Ethanol Exhibits Funding General anesthetic drugs Genetic Engineering Goals Hydrogen Bonding Ion Channel Ion Channel Protein Kinetics Laboratories Length Light Lipid Bilayers Maps Methods Modeling Molecular Movement Mutagenesis National Institute on Alcohol Abuse and Alcoholism Neurons Organism Pharmaceutical Preparations Physiological Positioning Attribute Potassium Channel Probability Property Proteins Research Personnel Resistance Role Scanning Series Site Solutions Structure Study Subject System Tail Testing Therapeutic Transplantation Voltage-Gated Potassium Channel Work alcohol effect alcohol sensitivity base insight interest member mutant patch clamp programs protein purification protein structure reconstitution response structural biology synthetic peptide three dimensional structure

项目摘要

DESCRIPTION (provided by applicant): The long-term goal of this project is to determine the structural basis of the functional modulation of brain ion channels by alcohol. The subject of this study is the neuronal Shaw2 potassium channel, which is directly inhibited by pharmacologically relevant concentrations of ethanol and other short-chain members of the homologous series of 1-alkanols. Guided by previous results from this study, the central hypothesis of the project states that the activation gate of the Shaw2 channel contributes an amphipathic protein-protein interface that constitutes the alcohol binding site. This project focuses on investigating the activation gate as the locus of the interactions that control the alcohol-induced responses of the Shaw2 channel. The specific aims of the project are: 1) To investigate the specific structural determinants of alcohol binding in the S4-S5 loop of the Shaw2 potassium channel. 2) To investigate the structural features that govern the alcohol-induced responses of the Shaw2 potassium channel in the C-terminal section of the S6 segment. 3) To develop the prokaryotic KvAP channel as a model to explore the achitecture of the alcohol binding site in the Shaw2 potassium channel. The first two aims systematically combine recombinant DNA technology, electrophysiology and biophysical analyses to map the alcohol site in the Shaw2 channel. In light of the recently solved crystal structure of the KvAP channel, the last aim seeks the potential application of structural biology to determine the achitecture of an engineered alcohol site. A detailed map of the physiologically relevant amphipathic interfaces that bind alcohol in ion channels is the first step toward understanding acute alcohol intoxication at the atomic level and targeting these sites for therapeutic applications.

描述（由申请人提供）：该项目的长期目标是确定酒精对脑离子通道功能调节的结构基础。本研究的主题是神经元 Shaw2 钾通道，该通道可直接被药理学相关浓度的乙醇和 1-烷醇同源系列的其他短链成员抑制。在本研究之前的结果的指导下，该项目的中心假设指出，Shaw2 通道的激活门贡献了构成酒精结合位点的两亲性蛋白质-蛋白质界面。该项目重点研究激活门作为控制 Shaw2 通道酒精诱导反应的相互作用轨迹。该项目的具体目标是： 1) 研究 Shaw2 钾通道 S4-S5 环中酒精结合的具体结构决定因素。 2) 研究控制 S6 片段 C 端部分 Shaw2 钾通道酒精诱导反应的结构特征。 3）以原核KvAP通道为模型，探索Shaw2钾通道中酒精结合位点的结构。前两个目标系统地结合重组 DNA 技术、电生理学和生物物理分析来绘制 Shaw2 通道中的酒精位点。鉴于最近解决的 KvAP 通道晶体结构，最后一个目标是寻求结构生物学的潜在应用来确定工程酒精位点的结构。在离子通道中结合酒精的生理相关两亲界面的详细图谱是在原子水平上了解急性酒精中毒并针对这些位点进行治疗应用的第一步。