3D Structure and Function of the Cys-loop Family of Ligand-gated Ion Channels

描述（由申请人提供）：Cys-loop受体是介导中枢和外周神经系统快速神经传递的五聚体配体门控离子通道（lgic），因其氨基末端细胞外配体结合域的保守二硫键而得名。这个整体膜蛋白超家族包括乙酰胆碱、血清素、γ -氨基丁酸、甘氨酸和几种无脊椎动物受体的离子电泳受体。这些亚家族中的每一个都包括具有独特配体结合和通道门控特性的受体亚型，这是由它们特定的亚基组成决定的。虽然受体亚型之间对配体的偏好和门控动力学存在很大差异，但在所有Cys-loop受体中，将小分子激动剂结合到约80埃远的通道开放的变构激活机制被认为是保守的。因此，对单个受体亚型的研究将为亚型选择疗法的设计提供信息，同时对整个Cys-loop家族的受体功能机制具有更广泛的意义。目前，还没有这个受体家族的原子分辨率结构信息；大多数结构信息是基于4埃电子显微镜结构和自然发生的可溶性细胞外结构域的晶体学结构。本提案的总体目标是在高分辨率下确定Cys-loop受体家族成员的3D结构。这一目标将通过在异源表达系统中检查已知家族的代表性成员来确定最佳的结晶候选者来实现；然后，一个ys-loop受体将被结晶并确定其结构。结构研究将由功能分析补充，测试关于受体状态转变机制的新的基于结构的假设。研究结果将对该家族和该亚型具有广泛的意义。拟议的研究将为神经退行性疾病（如阿尔茨海默病、帕金森病和精神分裂症）的主要治疗靶点提供见解。研究结果还将提高我们为这些疾病设计选择性治疗方法的能力，从而限制有害的副作用。

DESCRIPTION (provided by applicant): Cys-loop receptors are pentameric ligand-gated ion channels (LGICs) that mediate fast neurotransmission in the central and peripheral nervous systems, and are so named due to a conserved disulfide linkage in their amino terminal, extracellular ligand binding domains. This superfamily of integral membrane proteins includes the ionophoretic receptors for acetylcholine, serotonin, gamma-aminobutyric acid, glycine, and several invertebrate receptors. Each of these subfamilies comprises receptor subtypes with unique ligand binding and channel gating properties as determined by their specific subunit composition. While there is great variation in the preferences for ligands and gating kinetics between receptor subtypes, the allosteric activation mechanism that communicates binding of small molecule agonists into channel opening some 80-angstroms distant is believed to be conserved among all Cys-loop receptors. Hence, studies on an individual receptor subtype will be informative in terms of design of subtype-selective therapeutics, while simultaneously having broader implications for mechanisms of receptor function for the entire Cys-loop family. Currently, no atomic-resolution structural information exists for this family of receptors; most structural information is based on a 4-angstrom electron microscopy structure and crystallographic structures of a naturally-occurring soluble extracellular domain. The overall goal of this proposal is to determine the 3D structure of a Cys-loop receptor family member at high resolution. This goal will be accomplished by examining representative members from the known families in heterologous expression systems to determine the best candidates for crystallization; a Cys-loop receptor will then be crystallized and its structure determined. The structural studies will be complemented by functional assays that test new structure-based hypothesis regarding mechanisms of state transitions in the receptor. The results will have broad implications for the family in general and the subtype specifically. The proposed studies will provide insight into the major therapeutic targets for neurodegenerative disorders such as Alzheimer's and Parkinson's diseases, and schizophrenia. The results will also improve our ability to design selective therapeutics for these illnesses, thereby limiting deleterious side effects.