Structural biology of neurotransmitter ion channels

项目摘要 电可兴奋细胞分布在人体各处，在发育和发育过程中发挥着关键作用。 所有器官的功能，对神经的产生和正常活动特别重要 系统。配基门控离子通道(LGICs)是一种存在于细胞表面的多聚体完整膜蛋白。 可兴奋细胞的细胞膜和转导小分子配体的结合--所谓的化学 传递器-进入穿过细胞膜的离子传导通道的开口，从而传递 化学物质转化为电信号的梯度，电信号随后传播到相邻的细胞。因为 LGIC在可兴奋细胞的活动中扮演的中心角色，因为可兴奋细胞支持 许多器官和神经系统的功能，LGIC是大量治疗的靶点 药物，包括镇静剂、抗惊厥剂和抗恶心药物。此外，多个 疾病和障碍与LGICs的功能障碍有关。五聚体Cys环受体和 ATP门控的P2X受体是LGIC中最重要和最广泛的两类，被广泛应用 在人体内表达，是治疗剂的长期靶标。此外，这些 几十年来，两类LGIC一直是密集的生物物理和生化研究的对象。 尽管LGIC在医学和基础科学中具有重要的意义和历史意义，但对其研究 事实证明，3D结构很难实现。本申请中提出的研究将应用新技术和 显著促进两种环状受体亚型结构研究的试剂-甘氨酸和 GABAA受体-和三聚体、ATP门控的P2X受体。甘氨酸和GABAA受体尤其 重要的是，它们分别设定了外周和中枢神经系统的抑制性音调， 是一大批高效治疗药物的靶标。此外，P2X受体是 治疗疼痛、炎症和咳嗽的新靶点。尽管甘氨酸和 GABAA受体，以及P2X受体，在人类生物学和疾病中，我们还不了解，在 分子细节，受体如何被激动剂激活，如何被拮抗剂抑制，以及如何被 结合在新的位置的小分子的作用。此外，我们不了解这些LGIC是如何过渡的 从激动剂结合的开放状态到激动剂结合的脱敏或不活跃状态。使用最尖端的单一 粒子冷冻电子显微镜方法和X射线衍射方法，以及新的试剂和 方法，我们将定义甘氨酸和GABAA受体的活性机制，以及 人P2X3受体。这些进步反过来又为小说的发展奠定了基础 并更深入地了解原子结构和原子结构之间的关系。 Cys-loop和P2X受体家族的分子功能。

Project Summary Electrically excitable cells are spread throughout the human body, playing crucial roles in the development and function of all organs, and are particularly fundamental to the genesis and normal activity of the nervous system. Ligand-gated ion channels (LGICs) are multimeric integral membrane proteins that reside on the cellular membranes of excitable cells and transduce the binding of small molecule ligands – so called chemical transmitters – into the opening of an ion conducting channel across the cell membrane, thereby transducing a gradient of a chemical into an electrical signal that, in turn, is propagated to adjacent cells. Because of the central roles that LGICs play in the activities of excitable cells, and because excitable cells underpin the function of many organs and of the nervous system, LGICs are the targets of a large number of therapeutic drugs, including sedatives, anticonvulsive agents, and anti nausea medications. Furthermore, multiple diseases and disorders are associated with the dysfunction of LGICs. Pentameric Cys-loop receptors and ATP-gated P2X receptors are two of the most important and widespread classes of LGICs, are widely expressed throughout the human body and are long standing targets of therapeutic agents. Moreover, these two classes of LGICs have been the subject of intensive biophysical and biochemical studies for decades. Despite the significance and historical importance of LGICs in medicine and basic science, the study of their 3D structures has proven difficult. The research proposed in this application will apply new technologies and reagents to dramatically advance the structural study of two Cys-loop receptor subtypes – the glycine and GABAA receptor – and of trimeric, ATP-gated P2X receptors. Glycine and GABAA receptors are particularly important because they set the inhibitory `tone' of the peripheral and central nervous systems, respectively, and are the targets of a large number of highly efficacious therapeutic agents. Moreover, P2X receptors are emerging targets for the treatment of pain, inflammation and cough. Despite the prominence of glycine and GABAA receptors, as well as P2X receptors, in human biology and disease, we do not understand, in molecular detail, how the receptors are activated by agonists, inhibited by antagonists, and modulated by the action of small molecules binding at novel sites. Moreover, we do not understand how these LGICs transition from agonist-bound open states to agonist-bound desensitized or inactive states. Using cutting edge single particle cryo-electron microscopy methods and x-ray diffraction approaches, together with novel reagents and methods, we will define mechanisms for the activity of the glycine and GABAA receptors, as well as for the human P2X3 receptor. These advances, in turn, will provide a foundation for the development of novel therapeutic agents and for a deeper understanding of the relationships between atomic structure and molecular function in the Cys-loop and P2X receptor families.