Properties of Specific Alcohol Binding Sites

特定醇结合位点的特性

• 批准号: 7644537
• 负责人: JAMES Robert TRUDELL
• 金额: $ 30.5万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-07-01 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7644537
• 关键词: 3-Dimensional; Acetylcholine; Affect; Agonist; Agreement; Alcohol abuse; Alcoholism; Alcohols; Amino Acids; Arts; Binding; Binding Sites; Computer Simulation; Cysteine; Data; Dependence; Dimensions; Disulfides; Dodecanol; Engineering; Gated Ion Channel; Glycine; Goals; Health; Heptanol; In Vitro; Intoxication; Ion Channel; Ion Channel Protein; Knowledge; Laboratories; Ligands; Measures; Mediating; Membrane; Methods; Modeling; Modification; Molecular; Molecular Models; Mutation; Neurons; Oocytes; Property; Proteins; Reagent; Research Personnel; Rotation; Series; Serotonin Receptors 5-HT-3; Site; Solutions; Structure; Sulfhydryl Compounds; System; Testing; Torpedo; Transmembrane Domain; alcohol effect; alpha helix; analog; base; computer generated; crosslink; design; gamma-Aminobutyric Acid; in vivo; interfacial; methanethiosulfonate; molecular modeling; programs; propanethiol; receptor

DESCRIPTION (provided by applicant): Alcohol abuse and alcoholism are major health problems. It is likely that a solution to these problems will require an understanding of the effects of alcohol on neuronal ion channel proteins. Specific binding sites for alcohols have been described in the transmembrane (TM) domain of the superfamily of ligand-gated ion channels (LGIC's) that includes glycine (GlyRa1), GABA, nicotinic acetylcholine, and 5-HT3 receptors. Our global hypothesis is that alcohols bind within cavities that are bounded by TM segments of these receptors and preferentially stabilize specific channel substates. Our goal is to define the properties of those sites that mediate binding and efficacy of alcohol and alcohol antagonists in GlyRa1. These binding sites may provide a common motif for binding of alcohols within other classes of ion channels and other important proteins. We will build computational models of binding sites in GlyRa1 and design specific site-directed mutations to test these hypothesis. These mutations will be constructed and tested by our collaborator, Dr. R. A. Harris. Aim 1. We will test the hypothesis that amino acid residues from all four transmembrane helices of GlyRa1 contribute to a binding site for alcohols. We will develop computational models to delineate the dimensions of these sites. We will use these models to predict where covalent binding of alkyl methanethiosulfonate (MTS) reagents would mimic alcohol binding. The predictions will be tested in the Harris laboratory by expressing GlyRa1 containing site-directed cysteine substitutions in oocytes. They will apply MTS reagents to the oocytes and measure the effects on glycine-induced currents. Aim 2. We will test the hypothesis that double site directed cysteine mutations can clarify the refined tertiary structure of the GlyRa1 and distinguish our model from one based on the torpedo nAChR. While the overall structure of our GlyRa1 model and the nAChR model of Unwin are in global agreement, there are important differences in the orientation of transmembrane helices and residues bounding a possible alcohol- binding site. The Harris laboratory will test these predictions by crosslinking site-directed di-cysteines. In summary, knowledge of alcohol binding sites in GlyRa1 will increase our understanding of alcohol action in LGIC's. The results may reveal general motifs for action in other systems affected by alcohol and aid in the design of alcohol antagonists.

描述（由申请人提供）：酗酒和酗酒是主要的健康问题。解决这些问题可能需要了解酒精对神经元离子通道蛋白的影响。醇的特异性结合位点已在配体门控离子通道 (LGIC) 超家族的跨膜 (TM) 结构域中得到描述，该超家族包括甘氨酸 (GlyRa1)、GABA、烟碱乙酰胆碱和 5-HT3 受体。我们的总体假设是，醇类结合在以这些受体的 TM 片段为界的空腔内，并优先稳定特定的通道亚状态。我们的目标是确定 GlyRa1 中介导酒精和酒精拮抗剂结合和功效的位点的特性。这些结合位点可以为其他类别的离子通道和其他重要蛋白质内的醇的结合提供共同的基序。我们将建立 GlyRa1 结合位点的计算模型，并设计特定的定点突变来检验这些假设。这些突变将由我们的合作者 R.A. Harris 博士构建和测试。目标 1. 我们将检验以下假设：GlyRa1 的所有四个跨膜螺旋的氨基酸残基均有助于醇的结合位点。我们将开发计算模型来描绘这些地点的尺寸。我们将使用这些模型来预测烷基甲硫代磺酸盐 (MTS) 试剂的共价结合将在哪里模拟酒精结合。这些预测将在 Harris 实验室中通过在卵母细胞中表达含有定点半胱氨酸取代的 GlyRa1 进行测试。他们将 MTS 试剂应用于卵母细胞并测量对甘氨酸诱导电流的影响。目标 2. 我们将测试以下假设：双位点定向半胱氨酸突变可以澄清 GlyRa1 的精细三级结构，并将我们的模型与基于鱼雷 nAChR 的模型区分开来。虽然我们的 GlyRa1 模型​​和 Unwin 的 nAChR 模型的整体结构在全球范围内一致，但跨膜螺旋的方向和限制可能的醇结合位点的残基存在重要差异。哈里斯实验室将通过交联定点二半胱氨酸来测试这些预测。总之，对 GlyRa1 中酒精结合位点的了解将增加我们对 LGIC 中酒精作用的理解。结果可能揭示受酒精影响的其他系统中的一般作用模式，并有助于设计酒精拮抗剂。

项目成果

Microsecond simulations indicate that ethanol binds between subunits and could stabilize an open-state model of a glycine receptor.

微秒模拟表明，乙醇在亚基之间结合，可以稳定甘氨酸受体的开放状态模型。

• DOI: 10.1016/j.bpj.2011.02.032
• 发表时间: 2011
• 期刊: Biophysical journal
• 影响因子: 3.4
• 作者: Murail,Samuel;Wallner,Björn;Trudell,JamesR;Bertaccini,Edward;Lindahl,Erik
• 通讯作者: Lindahl,Erik

Ethanol inhibition of constitutively open N-methyl-D-aspartate receptors.

乙醇对开放的 N-甲基-D-天冬氨酸受体的抑制。

• DOI: 10.1124/jpet.111.187179
• 发表时间: 2012
• 期刊: The Journal of pharmacology and experimental therapeutics
• 作者: Xu,Minfu;Smothers,CThetford;Trudell,James;Woodward,JohnJ
• 通讯作者: Woodward,JohnJ

Ethanol Modulation is Quantitatively Determined by the Transmembrane Domain of Human α1 Glycine Receptors.

乙醇调节是由人类α1 甘氨酸受体的跨膜域定量确定的。

• DOI: 10.1111/acer.12735
• 发表时间: 2015
• 期刊: Alcoholism, clinical and experimental research
• 作者: Horani,Suzzane;Stater,EvanP;Corringer,Pierre-Jean;Trudell,JamesR;Harris,RAdron;Howard,RebeccaJ
• 通讯作者: Howard,RebeccaJ

Comparative modeling of a GABAA alpha1 receptor using three crystal structures as templates.

使用三种晶体结构作为模板对 GABAA α1 受体进行比较建模。

• DOI: 10.1016/j.jmgm.2004.03.004
• 发表时间: 2004
• 期刊: Journal of molecular graphics & modelling
• 影响因子: 2.9
• 作者: Trudell,JR;Bertaccini,E
• 通讯作者: Bertaccini,E

Tryptophan 46 is a site for ethanol and ivermectin action in P2X4 receptors.

色氨酸 46 是 P2X4 受体中乙醇和伊维菌素作用的位点。

• DOI: 10.1007/s11302-013-9373-4
• 发表时间: 2013
• 期刊: Purinergic signalling
• 影响因子: 3.5
• 作者: Popova,Maya;Trudell,James;Li,Kaixun;Alkana,Ronald;Davies,Daryl;Asatryan,Liana
• 通讯作者: Asatryan,Liana