Correlations: Structure-Dynamics-Function in Channels

相关性：通道中的结构-动力学-功能

基本信息

批准号：
8711160
负责人：
TIMOTHY A CROSS
金额：
$ 46.47万
依托单位：
FLORIDA STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1987
资助国家：
美国
起止时间：
1987-01-01 至 2015-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8711160
关键词：
Acids Affect Amantadine Antidotes Antiviral Agents Attention Binding Binding Sites Biological Assay Biophysics Chemicals Coupled Crystallography Data Drug Binding Site Drug Targeting Drug resistance Environment Genes Health Heart Heterogeneity Histidine Hydrophobicity Influenza Influenza A Virus, H1N1 Subtype Ion Channel Knowledge Length Lipid Bilayers Lipids Liposomes Location M2 protein Magic Membrane Membrane Proteins Methods Modeling Molecular Molecular Conformation Motion Mutation NMR Spectroscopy Oocytes Pharmaceutical Preparations Positioning Attribute Proteins Protons Quantum Mechanics Research Resolution Rimantadine Role Sampling Shapes Solutions Structure Transmembrane Domain Vertebral column Viral Virus Water chemical property design insight meetings mimetics molecular dynamics molecular mechanics mutant novel pandemic disease pandemic influenza prophylactic protein structure restraint solid state nuclear magnetic resonance swine flu

项目摘要

DESCRIPTION (provided by applicant): M2 protein in the influenza A membrane coat is a known drug target. Unfortunately, the S31N mutation has caused widespread drug resistance in H3N2 since 2006 and in the recent H1N1 swine flu pandemic. Excellent progress has been made in understanding the H+ transport activity of this protein through characterizations of truncated versions of M2 that form partially functional transporters. At the heart of this tetrameric protein are four histidine residues that have been shown to have unique chemical properties responsible for the acid activation of the pore and suspected of being directly involved in the transporter mechanism for H+ conductance. Drug binding in the pore dramatically influences the pKas of the histidine residues, displaces waters that facilitate helix kinking, and induces the closure of the Val27 secondary gate. Studies of the M2 truncates have been complicated by membrane mimetic environments used for structural characterization. The comparison of structures obtained by x-ray crystallography, solution NMR and solid state NMR spectroscopy clearly show that the environment has a substantial influence upon secondary, tertiary, and quaternary levels of protein structure. Here, structural, dynamical, and functional characterizations of the full-length protein will all be performed in bilayers having a lipid composition similar to that of the viral membrane. Liposome and oocyte assays of H+ conductance, magic angle spinning and aligned sample solid state NMR spectroscopy, and molecular dynamics simulations will be performed using nearly identical environments for the full-length M2 protein. High-resolution complete (backbone and sidechains) structural characterizations as a function of pH and in the presence and absence of drug will be achieved. This structural data will be coupled with conductance assays and molecular dynamics simulations to achieve a quantitative atomistic mechanism for selective proton conductance and acid activation. Similarly, data from the S31N mutant and in the presence and absence of amantadine will be combined to reach a detailed understanding of how the drug blocks conductance in the wild-type protein but not in the S31N mutant. Detailed functional understanding at this level is difficult to achieve for a membrane protein. Here, a proven interdisciplinary team will meet this challenge for a proven drug target that could once again be an important factor in the defense against a deadly influenza pandemic.

描述（由申请人提供）：流感中的M2蛋白膜外套是已知的药物靶标。不幸的是，自2006年和最近的H1N1猪流感大流行中，S31N突变引起了H3N2的广泛耐药性。通过构成部分功能转运蛋白的M2的截短版本的特征来理解该蛋白质的H+转运活性。该四聚体蛋白的核心是四个组氨酸残基，这些残基已被证明具有负责孔的酸激活的独特化学特性，并怀疑直接参与了用于H+电导的转运蛋白机制。孔中的药物结合极大地影响了组氨酸残基的PKA，置换了促进螺旋扭结的水，并诱导Val27二级门的闭合。 M2截断的研究使用于结构表征的膜模拟环境变得复杂。通过X射线晶体学，溶液NMR和固态NMR光谱获得的结构的比较清楚地表明，环境对蛋白质结构的二级，第三和第四纪水平具有重大影响。在这里，全长蛋白的结构，动力学和功能特征都将在具有类似于病毒膜的脂质组成的双层中进行。 H+电导，魔法角度旋转和样品固态NMR光谱和分子动力学模拟的脂质体和卵母细胞测定将使用几乎相同的环境进行全长M2蛋白进行。将实现高分辨率完整（骨干和侧链）结构特征，并在存在和不存在药物的情况下进行结构特征。该结构数据将与电导分析和分子动力学模拟相结合，以实现选择性质子电导和酸激活的定量原子机制。同样，将从S31N突变体以及在存在和不存在的情况下组合使用的数据，以详细了解野生型蛋白中的药物的电导方式，而不是在S31N突变体中。对于膜蛋白，很难在此水平上详细的功能理解。在这里，一个经过验证的跨学科团队将面对这一挑战，这可能再次成为防御致命流感大流行的重要因素。