Probing allosteric surfaces of NMDA receptors

描述（由申请人提供）：许多内源性和合成配体调节NMDA受体活性，是许多神经系统疾病治疗干预的潜在候选者。迄今为止，控制NMDA受体介导的通量的经验尝试在临床中仅取得了适度的成功，主要是由于对NMDA受体活性变构控制的机制以及这些活动在脑生理和病理中所起的具体作用的理解不足。在之前的资助期内，目标是描述内源性调节剂（质子、锌/伊芬普罗地尔、甘氨酸）影响NMDA受体门控动力学的机制，从而控制与突触信号传导相关的宏观反应。在下一个资助期，目标是描绘构成NMDA受体激活的细胞内蛋白质运动。一般的方法是利用最近解决的GluA2四聚体受体的原子分辨率结构，NMDA受体同源物和我们在NMDA受体门控调制方面不断增长的专业知识。我们将引入突变来干扰（增加或减少）NMDA受体结构模块的相对迁移率，并将通过单分子信号的动力学分析来描述反应机制的伴随变化。此外，我们将探索受限制或增强内部运动的受体获得宏观反应的时间过程，以更好地了解特定的结构特征如何支持NMDA受体在脑生理和病理中发挥的独特生物学功能。总的来说，这项工作将提供有关NMDA受体激活的结构相关的关键信息，并将整合目前孤立的门控结构和动力学模型。鉴于谷氨酸受体介导超过90%的大脑兴奋传递，NMDA受体对许多基本的大脑功能至关重要，本实验产生的知识可能对神经传递和神经调节领域产生广泛影响。

DESCRIPTION (provided by applicant): Numerous endogenous and synthetic ligands modulate NMDA receptor activities and are potential candidates for therapeutic intervention in a number of neurologic disorders. To date, empirical attempts to control en masse NMDA receptor-mediated fluxes have had only modest success in the clinic, mainly due to inadequate understanding of the mechanisms governing the allosteric control of NMDA receptor activities and of the specific roles played by these activities in brain physiology and pathology. Over the previous funding period the objective has been to delineate the mechanisms by which endogenous modulators (protons, zinc/ifenprodil, glycine) affect NMDA receptor gating dynamics and thus control the macroscopic response relevant to synaptic signaling. Over the next funding period the objective is to delineate the intracellular protein motions that constitute the NMDA receptor activation. The general approach is to capitalize on the recently solved atomic-resolution structure for a GluA2 tetrameric receptor, a NMDA receptor homologue and our growing expertise on NMDA receptor gating modulation. We will introduce mutations to perturb (increase and decrease) the relative mobility of NMDA receptor structural modules and will delineate the accompanying changes in reaction mechanism by kinetic analyses of single-molecule signals. Further, we will explore the time course of macroscopic responses obtained from receptors with restricted or enhanced internal motions to better understand how specific structural features support the unique biological functions played by NMDA receptors in brain physiology and pathology. Overall this work will provide critical information about structural correlates of NMDA receptor activation and will integrate the currently isolated structural and kinetic models of gating. Given that glutamate receptors mediate more than 90% of excitatory transmission in brain and NMDA receptors are critical to many fundamental brain functions, knowledge generated by the proposed experiments is likely to have wide impact on the fields of neurotransmission and neuromodulation.