Defining Alcohol Binding Sites in Ligand-Gated Ion Channels

描述（由申请人提供）：酒精滥用和酒精中毒是影响1700多万人的重大健康问题，每年造成近2000亿美元的损失。解决这些问题的方法很可能是更好地了解酒精对神经元离子通道和调节它们的蛋白质的影响。拟议研究的目标将是在原子水平上理解醇结合位点的特性的重要一步。我们相信，这些知识对于未来设计和选择能够减少酒精引起的渴望或成瘾的药物至关重要。具体来说，我们将研究配体门控离子通道（lgic）中醇结合的位点，包括GABAaRs和GlyRs。我们的同源性建模和实验方法将提供GABAaRs的三维可视化，以增加我们对酒精作用的理解。这种创新的方法将尖端的计算和神经科学技术与分子生物学相结合。我们期望我们的结果将对神经系统其他重要受体中更广泛的酒精结合位点产生重大影响。我们的方法通过三个具体目标关注醇结合位点的三个方面：醇结合位点在哪里；亚基内与亚基间

DESCRIPTION (provided by applicant): Alcohol abuse and alcoholism are significant health problems that affect over 17 million people and cost nearly $200 billion annually. It is likely tha a solution to these problems will result from a better understanding of alcohol effects on neuronal ion channels and the proteins that modulate them. The goal of the proposed research will be a significant step towards understanding the properties of alcohol-binding sites at an atomic level. We believe this knowledge will be essential for future design and selection of drugs that could reduce craving or addiction induced by alcohol. Specifically, we will study the sites fo alcohol binding in ligand- gated ion channels (LGICs), which include GABAaRs and GlyRs. Our homology modeling and experimental methods will provide 3-dimensional visualization of GABAaRs to increase our understanding of alcohol's action. This innovative approach combines cutting-edge computational and neuroscience techniques with molecular biology. We expect our results will have a significant impact on the broader class of alcohol- binding sites in other important receptors of the nervous system. Our Approach focuses on three aspects of alcohol-binding sites via three Specific Aims: Where are alcohol-binding sites; intra-subunit versus inter- subunit in LGICs (Aim 1), which specific residues or segments in LGICs mediate the effect of alcohol binding at these sites (Aims 1 and 2), and how do these sites modulate ligand binding (Aims 2 and 3). In Aim 1, Trudell and Bertaccini will build computational models of alcohol-binding sites in GABA receptors and design site-directed mutations to test the models. Harris and Howard, under a subcontract to the University of Texas, Austin, will test the function of these mutated receptors. We will iteratively refine the models the Trudell group will use the models to predict the effects of mutations; the Harris group will test if the models are consistent with experimental data; and the Trudell group will then modify the models to fit the new data. They will address this controversial question: What is the most important alcohol effect site in GABAaR? Is it Intra-subunit or Inter-subunit? They will also test the hypothesis that the GABAaR TM3 helix must rotate during activation in order to incorporate all recent experimental data. In Aim 2, the Trudell and Harris laboratories will recreate the alcohol-binding site from GABAaRs in the homologous but natively EtOH insensitive ion channel, GLIC, by determining which residues are specific to EtOH binding in GABAR and mutating these into their corresponding homologous positions within GLIC. In Aim 3, Trudell and Bertaccini will use three docking programs to investigate binding of alcohol analogs. Our investigators have proven accomplishment in alcohol research and possess the resources necessary to accomplish our Aims. Our proposal is responsive to both the NIAAA initiative in computational neuroscience and the NIH Roadmap: Bioinformatics and Computational Biology. These significant studies will provide essential knowledge needed to design alcohol-binding antagonists which could revolutionize treatment for alcohol abuse and dependence.