Molecular Mechanisms of Glutamate Receptor Channels

描述（由申请人提供）： 我们拟研究谷氨酸受体通道（GluR）门控。GluR介导大脑中大量的兴奋性神经传递，并且恢复由于GluR激活而下降的离子梯度所需的能量代表生物体总能量预算的显著部分。NMDA受体是GluR的一种亚型，产生缓慢的去极化反应，并参与突触发育和可塑性。NMDA受体门控作为三个不同的构象步骤发生，并且缓慢的突触衰减在很大程度上是由开放状态的“陷阱”引起的，所述“陷阱”维持电流流动并且保持受体不采用激动剂可以解离的构象。此外，单个NMDA受体可以产生三种不同的门控动力学模式，这些模式是由开放构象的相对稳定性波动引起的。我们建议通过测量扰动（激动剂或侧链突变的变化）对三个门控反应的详细单通道动力学的影响来探索NMDA受体门控的能量景观。我们的方法是使用线性自由能分析来揭示分子运动的顺序和组织，当蛋白质在“封闭”和“开放”之间穿越构象景观时发生。此外，我们建议探测第二类重要的GluRs，AMPA受体的门控反应。我们的总体目标是了解构成门控GluRs的分子事件，并将此信息的背景下，门控其他突触受体，离子通道和变构蛋白。因此，我们将更好地理解递质、药物和其他配体塑造表达这些重要受体通道蛋白的神经元的细胞反应的机制。

DESCRIPTION (provided by applicant): We propose to study glutamate receptor-channel (GluR) gating. GluRs mediate the bulk of excitatory neurotransmission in the brain, and the energy required to restore the ionic gradients that run down as a consequence of GluR activation represents a significant fraction of the total energy budget of the organism. NMDA receptors are a subtype of GluR that generate a slow, depolarizing response and that participate in synaptic development and plasticity. NMDA receptor gating occurs as three distinct conformational steps, and the slow synaptic decay arises, to a large extent, from an open state 'trap' that maintains current flow and that keeps the receptor from adopting a conformation from which agonists can dissociate. Moreover, an individual NMDA receptor can generate three distinct gating kinetic modes that arise from fluctuations in the relative stability of the open conformations. We propose to explore the energy landscape for NMDA receptor gating by measuring the effects of perturbations (changes in agonists or mutation of sidechains) on the detailed, single-channel kinetics of the three gating reactions. Our approach is to use linear free energy analysis to reveal the order and organization of molecular motions that occur as the protein traverses the conformational landscape between 'closed' and 'open'. In addition, we propose to probe the gating reaction(s) for a second important class of GluRs, AMPA receptors. Our overall objective is to understand the molecular events that constitute gating of GluRs and to place this information in the context of gating of other synaptic receptors, ion channels, and allosteric proteins. Accordingly, we will better understand the mechanisms by which transmitters, drugs, and other ligands shape the cellular responses of neurons that express these important receptor-channel proteins.