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中兴奋的传播， 最终控制生理功能，如运动和认知。它们被分为四类 亚家族：N-甲基-D-天冬氨酸（NMDA），N-氨基-5-甲基-3-羟基-4-异恶唑丙酸酯（AMPA）， 红藻氨酸盐和δ受体。所有四种亚型在拓扑结构上都是相似的，包括四个蛋白质亚基， 作为二聚体的二聚体。然而，它们的门控特性和机制是独特的，反映了 在突触传递中的不同作用。我实验室的目标是了解 这些亚型的功能微调。使用多管齐下的方法，结合生物化学， 电生理学，荧光和计算方法，我们已经确定了构象 这些转换对于介导AMPA和NMDA谷氨酸受体的激活和抑制至关重要。 然而，直接了解蛋白质控制的一部分的这种结构变化如何改变， 其他部分以及它们如何决定功能特性仍然是未知的。我们建议开发和使用 单分子荧光共振能量转移（FRET）绘制结构-功能 每个谷氨酸受体亚型的不同片段之间的相关性。我们还将集中力量， 了解研究较少的红藻氨酸和δ亚型。具体来说，我们将研究 激动剂、离子和辅助剂对丰富形式的红藻氨酸受体的潜在激活和调节 proteins.此外，我们有初步的数据表明，δ受体功能依赖于连接 与跨突触蛋白复合物，因此，我们将研究δ受体的结构和功能， 突触结合伴侣的背景。总之，这些研究将把我们基本的单分子 在生理学背景下的研究，并最终使我们能够了解功能差异之间 突触和非突触谷氨酸受体。

PROJECT SUMMARY/ABSTRACT Glutamate receptors mediate the spread of excitation in the mammalian central nervous system, and ultimately control physiological functions such as movement and cognition. They are classified into four subfamilies: N-methyl-D-aspartate (NMDA), -amino-5-methyl-3-hydroxy-4-isoxazole propionate (AMPA), kainate, and delta receptors. All four subtypes are topologically similar, comprising four protein subunits held together as a dimer-of-dimers. However, their gating characteristics and mechanisms are unique, reflecting distinct roles in synaptic transmission. The goal of my laboratory is to understand the mechanisms that underlie the functional fine-tuning of each of these subtypes. Using a multipronged approach that combines biochemical, electrophysiological, fluorescence, and computational methods, we have already identified conformational transitions that are critical for mediating activation and inhibition of AMPA and NMDA glutamate receptors. However, a direct understanding of how such structural changes in one part of the protein control changes in other parts, and how they dictate functional properties, remains unknown. We propose to develop and use multicolor single-molecule fluorescence resonance energy transfer (FRET) to draw structure–function correlations between different segments of each glutamate receptor subtype. We will also focus our efforts on understanding the less well-studied kainate and delta subtypes. Specifically, we will investigate the mechanisms underlying activation and modulation of an abundant form of the kainate receptor by agonists, ions, and auxiliary proteins. In addition, we have preliminary data showing that delta receptor function is dependent on connections with trans-synaptic protein complexes, therefore we will study the structure and function of delta receptors in the context of synaptic binding partners. Together, these investigations will place our fundamental single-molecule investigations in a physiological context, and eventually allow us to understand functional differences between synaptic and non-synaptic glutamate receptors.