Structure, function and dynamics of a glutamate receptor

嗜冷谷氨酸受体控制着多种正常的神经元过程，包括学习和 记忆。此外，这些重要的神经递质受体的激活参与了许多 神经退行性疾病，特别是中风和癫痫。因此，针对谷氨酸的药物 受体将具有相当大的治疗价值。对跨膜拓扑的分析导致了 实现每个亚基由一系列模块组成，与谷氨酸结合的模块可以是 在细菌中以可溶性蛋白质的形式产生(S1S2结构域)。S1S2结构域结合激动剂和 与完整受体具有大致相同亲和力的拮抗剂，是一种极好的系统 用于研究结合结构域。GluR2亚基的S1S2结构域的晶体结构 由E.Gouaux和合作者解决。我们已经使用核磁共振波谱来表征动态 GluR2 S1S2域的属性。不幸的是，GluR2受体的功能很难研究 由于其快速的动力学和低的通道电导。这项建议的目的是延长 形成更适宜通道的谷氨酸受体亚单位(GluRS)的结构和动力学研究 进行详细的动力学测量。核磁共振和荧光研究将表征该化合物的结构和 在一系列完全和部分激动剂存在的情况下的动力学。如晶体结构和 同源模型，S1S2结构域是一种双体结构，在配体结合时关闭。我们的假设 通道电导是由单个亚基中的波瓣闭合来调制的，或者大的- 尺度构象动力学或叶内动力学有助于电导水平的变化。 明渠暴发中的开启和关闭与内部动力学具有相同的时间尺度 S1S2域。将确定同构体GluRS的单通道电导和速率常数 同一系列激动剂中的受体-通道，以确定原子水平相互作用是否存在明显差异 激动剂和受体构象之间的差异影响通道动力学。内部的比较 具有通道的单通道属性的动力学应该允许我们确定这些动力学是否 发挥对离子通道功能的调控作用。最后一个目标将包括核磁共振和单声道录音 从突变的S1S2结构域确定叶间界面在激动剂亲和力控制中的作用 和药效。尽管同源GLURS受体通道可能仅限于少数 神经元，GLUR2/GLUR3异构体受体通道可能代表着大量的 突触后的AMPA受体在新皮质。这一结果将有助于揭示一种新的结合部位 重要的谷氨酸受体亚单位，为进一步的药物开发提供必要的信息。

lonotropic glutamate receptors control a variety of normal neuronal processes including learning and memory. In addition, activation of these important neurotransmitter receptors is involved in a number of neurodegenerative diseases, notably stroke and epilepsy. Thus, drugs targeted toward glutamate receptors would be of considerable therapeutic value. Analysis of the transmembrane topology led to the realization that each subunit is made of a series of modules, and the module that binds glutamate can be produced in bacteria as a soluble protein (S1S2 domain). The S1S2 domain binds agonists and antagonists with approximately the same affinity as the intact receptor and serves as an excellent system for studying the binding domain. The crystal structure of the S1S2 domain of the GluR2 subunit has been solved by E. Gouaux and collaborators. We have used NMR spectroscopy to characterize the dynamic properties of the GluR2 S1S2 domain. Unfortunately, the function of the GluR2 receptor is difficult to study due to its rapid kinetics and low channel conductance. The purpose of this proposal is to extend the structural and dynamic studies to a glutamate receptor subunit (GluRS) that forms channels more suitable for detailed kinetic measurements. NMR and fluorescence studies will characterize the structure and dynamics in the presence of a series of full and partial agonists. As shown by crystal structures and homology models, the S1S2 domain is a bilobe structure that closes upon ligand binding. Our hypothesis is that the channel conductance is modulated by lobe closure in individual subunits and that either large- scale conformational dynamics or intralobe dynamics contribute to changes in conductance levels. Openings and closings within an open channel burst are on the same timescale as the internal dynamics of the S1S2 domain. Single channel conductances and rate constants will be determined for homomeric GluRS receptor-channels in the same series of agonists to determine if clear differences in atomic level interactions between agonist and receptor conformation differentially affect channel kinetics. Comparison of the internal dynamics with the single channel properties of the channels should allow us to determine if these dynamics play a role in the control of ion channel function. The last aim will include NMR and single channel recordings from mutated S1S2 domains to determine the role of the interlobe interface in the control of agonist affinity and efficacy. Although homomeric GLURSreceptor-channels are likely to be confined to a small number of neurons, GLUR2/GLUR3 heteromeric receptor-channels may represent a significant number of postsynaptic AMPA receptors in the neocortex. The results will shed light on the binding site of an important glutamate receptor subunit and provide essential information for further drug development.