Structural studies of heteromeric nicotinic acetylcholine receptors

异聚烟碱乙酰胆碱受体的结构研究

• 批准号: 9155684
• 负责人: Ryan E Hibbs
• 金额: $ 36.43万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-07-01 至 2021-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9155684
• 关键词: Autonomic ganglion; Binding; Biochemical; Biological Assay; Biophysics; Brain; Cell Surface Receptors; Chemicals; Complication; Crystallization; Data; Data Set; Development; Electrophysiology (science); Family; Foundations; Future; Gated Ion Channel; Goals; Heterogeneity; Individual; Ions; Length; Ligands; Link; Measures; Mediating; Membrane Proteins; Mental Health; Mental disorders; Methods; Modeling; Molecular; Molecular Target; Nervous System Physiology; Neurodegenerative Disorders; Neurons; Neurotransmitter Receptor; Neurotransmitters; Nicotine; Nicotine Dependence; Nicotinic Receptors; Peripheral Nervous System; Pharmaceutical Preparations; Pharmacology; Play; Population; Preparation; Property; Proteins; Recombinants; Research; Resolution; Roentgen Rays; Structure; Sum; Testing; Therapeutic; abstracting; addiction; biochemical tools; design; design and construction; improved; insight; member; nervous system disorder; neurotransmission; patch clamp; radioligand; receptor; research study; stoichiometry; therapeutic target; transmission process

Abstract Neuronal nicotinic acetylcholine receptors are essential therapeutic targets for addiction, mental health and neurodegenerative disorders. These pentameric ligand-gated ion channels are the prototypical members of the Cys-loop receptor superfamily, which mediate fast neurotransmission throughout the central and peripheral nervous systems. Here we propose to determine high-resolution structures of two representative neuronal nicotinic receptor subtypes. Structural analysis of nicotinic receptors has been hampered by the challenges of recombinant expression of eukaryotic membrane proteins. These proteins typically express at low levels and are unstable after purification. Furthermore, most nicotinic receptors are obligate heteromers and in many cases these heteromers can assemble as pentamers with different ratios of subunits. This complication of mixed stoichiometry is present among all Cys-loop receptor families but is best characterized in the nicotinic receptors. The structural heterogeneity results in physiologically important, finely tuned pharmacological and channel properties. In Aim 1, we propose to develop methods for expression and purification of these receptors that can assemble in multiple stoichiometries. In Aims 2 and 3 we propose to apply these approaches to determine structures of two heteromeric nicotinic receptors in physiologically- relevant and functionally-distinct alternate stoichiometries. The individual receptor structures will provide key insights into the structural underpinnings of ion permeation and ligand recognition. Comparison of the structures of the two receptors, and of alternative stoichiometries of the same receptor, will provide a reliable structural foundation for understanding the distinctive biophysical and pharmacological properties of each receptor subunit combination.

抽象的 神经元烟碱乙酰胆碱受体是成瘾、心理健康的重要治疗靶点 和神经退行性疾病。这些五聚体配体门控离子通道是原型成员 Cys环受体超家族的成员，它介导整个中枢和神经系统的快速神经传递 周围神经系统。在这里，我们建议确定两个代表性的高分辨率结构 神经元烟碱受体亚型。烟碱受体的结构分析受到以下因素的阻碍： 真核膜蛋白重组表达的挑战。这些蛋白质通常表达于 含量低且纯化后不稳定。此外，大多数烟碱受体是专性异聚体 在许多情况下，这些异聚体可以组装成具有不同比例的亚基的五聚体。这 混合化学计量的并发症存在于所有 Cys 环受体家族中，但最好的特征是 烟碱受体。结构异质性导致生理上重要的、精细调整的 药理和通道特性。在目标 1 中，我们建议开发表达和表达的方法 纯化这些可以多种化学计量组装的受体。在目标 2 和 3 中，我们建议 应用这些方法来确定生理学中两种异聚烟碱受体的结构 相关且功能不同的替代化学计量。个体受体结构将提供关键 深入了解离子渗透和配体识别的结构基础。比较 两种受体的结构以及同一受体的替代化学计量的结构将提供可靠的 了解每种药物独特的生物物理和药理学特性的结构基础 受体亚基组合。