Dynamics of ligand gated ion channels

项目总结/摘要 谷氨酸受体介导哺乳动物中枢神经系统的兴奋性反应，并最终 控制运动和认知功能。谷氨酸受体根据其功能分为三个亚家族， 几种合成和天然激动剂的亲和力曲线：N-甲基-D-天冬氨酸（NMDA），N-氨基-5-甲基-3-甲基-N-氨基-D-天冬氨酸（NMDA）， 羟基-4-异恶唑丙酸酯（AMPA）和红藻氨酸受体。所有这三种亚型大致相似， 具有类似拓扑结构的二聚体的二聚体架构。然而，它们的门控特性和机制是 在突触传递中的独特作用。我实验室的工作重点是了解 这些亚型的微调机制。AMPA受体介导快速突触 传输和它们的门控特性也被辅助亚基（TARP）的存在所调制 通过翻译后修饰。我实验室的工作提供了深入了解 AMPA受体的激活和脱敏，使用生物物理和生物化学的组合， 方法.在这里，我们建议研究如何将这些机制转化为通过TARP进行的调制， 翻译后修饰如磷酸化。为此，我们将使用smFRET和 LRET确定动力学和构象变化，并验证这些结构-动力学变化 通过对由受体样交叉的生化操纵引起的变化的功能表征， 连接和突变。另一方面，NMDA受体介导Ca 2+渗透性长去极化 信号并通过小分子调节剂、磷酸化和相互作用伙伴（例如 作为钙调蛋白和α-辅肌动蛋白在细胞内羧基末端。smFRET和LRET研究， 我的实验室以及X射线和EM结构为构象提供了重要的见解 有序的细胞外结构域内的变化。然而，跨域的通信和角色 无序的C-末端结构域在很大程度上尚未探索。在这里，我们建议研究相互作用的作用 跨域控制受体的动力学和构象变化，以及调节剂的作用 和变化的C-末端结构域对这些相互作用和构象动力学。这些研究将 然后将其与功能性后果相关联，从而提供对结构-动力学途径的深入了解， 这些受体的激活、脱敏和调节。

PROJECT SUMMARY/ABSTRACT Glutamate receptors mediate excitatory responses in the mammalian central nervous system, and ultimately control motor and cognitive functions. Glutamate receptors are classified into three subfamilies based on affinity profiles for several synthetic and natural agonists: N-methyl-D-aspartate (NMDA), -amino-5-methyl-3- hydroxy-4-isoxazole propionate (AMPA) and kainate receptors. All three subtypes are broadly similar having a dimer of dimer architecture with similar topologies. However, their gating characteristics and mechanisms are unique resulting in unique roles in synaptic transmission. The work in my laboratory focuses on understanding the mechanisms underlying the fine-tuning of each of these subtypes. AMPA receptors mediate fast synaptic transmission and their gating properties are modulated by the presence of auxiliary subunits (TARPs) as well as through post-translational modifications. Work from my laboratory has provided insight into the mechanism of activation and desensitization in AMPA receptors using a combination of biophysical and biochemical methods. Here, we propose to study how these mechanisms can be translated to modulation by TARPs and post-translational modifications such as phosphorylation. For this we will use a combination of smFRET and LRET to determine the dynamics and conformational changes, and validate these structure-dynamic changes through functional characterization of changes elicited by biochemical manipulations of the receptor like cross linking and mutations. The NMDA receptors, on the other hand, mediate Ca2+ permeable long depolarizing signals and are modulated through small molecule modulators, phosphorylation and interacting partners such as calmodulin and alpha-actinin at the intracellular carboxy terminus. smFRET and LRET investigations from my laboratory as well as the X-ray and EM structures have provided significant insight into conformational changes within the ordered extracellular domains. However, the communication across domains and the role of the disordered C-terminal domain are largely unexplored. Here we propose to study the role of interactions across domains in controlling dynamics and conformational changes in the receptor, and effects of modulators and changes at the C-terminal domain on these interactions and conformational dynamics. These studies will then be correlated to functional consequences thus providing insight into the structure-dynamic pathway of activation, desensitization, and modulation in these receptors.