The Free Energy Landscape of GABA Binding to a Pentameric Ligand-Gated Ion Channel and Its Disruption by Mutations
The Free Energy Landscape of GABA Binding to a Pentameric Ligand-Gated Ion Channel and Its Disruption by Mutations
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DOI：
10.1021/acs.jctc.6b00303
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发表时间：
2016-07-01
影响因子：
5.5
通讯作者：
Molteni, Carla
中科院分区：
文献类型：
--
作者：
Comitani, Federico;Limongelli, Vittorio;Molteni, Carla
Cys-loop超家族的五聚体配体门控离子通道（plgic）是介导快速突触传递的重要神经受体。它们是由一种神经递质结合而激活的，但这一过程的细节尚不完全清楚。作为一种典型的pLGIC，我们选择了参与杀虫剂抗性的昆虫抗狄氏剂受体（RDL），并在原子水平上研究神经递质GABA与其胞外结构域的结合。我们通过多秒漏斗元动力学模拟实现了这一目标，该模拟利用漏斗形约束势来限制溶剂中的勘探，有效地增强了束缚态和非束缚态的采样。我们揭示了结合过程中的一系列事件，从GABA从溶剂中捕获到结合口袋中带电残基Arg111和Glu204之间的固定。我们描述了野生型RDL受体和两种突变形式中相关的自由能景观，其中关键残基Arg111和Glu204突变为Ala。实验上，这些突变产生了非功能性通道，这反映在由于基本相互作用的丧失而降低的配体结合亲和力上。我们还分析了关键环C的动力学行为，其打开允许GABA进入结合位点，关闭将配体锁定在蛋白质中。RDL受体与其他plgic具有相同的结构和功能特征；因此，我们的工作概述了一个有价值的方案来研究配体与plgic的结合，而不是传统的对接和分子动力学技术。
Pentameric ligand-gated ion channels (pLGICs) of the Cys-loop superfamily are important neuroreceptors that mediate fast synaptic transmission. They are activated by the binding of a neurotransmitter, but the details of this process are still not fully understood. As a prototypical pLGIC, here we choose the insect resistance to dieldrin (RDL) receptor involved in resistance to insecticides and investigate the binding of the neurotransmitter GABA to its extracellular domain at the atomistic level. We achieve this by means of mu-sec funnel-metadynamics simulations, which efficiently enhance the sampling of bound and unbound states by using a funnel-shaped restraining potential to limit the exploration in the solvent. We reveal the sequence of events in the binding process from the capture of GABA from the solvent to its pinning between the charged residues Arg111 and Glu204 in the binding pocket. We characterize the associated free energy landscapes in the wild-type RDL receptor and in two mutant forms, where the key residues Arg111 and Glu204 are mutated to Ala. Experimentally these mutations produce nonfunctional channels, which is reflected in the reduced ligand binding affinities due to the loss of essential interactions. We also analyze the dynamical behavior of the crucial loop C, whose opening allows the access of GABA to the binding site and closure locks the ligand into the protein. The RDL receptor shares structural and functional features with other pLGICs; hence, our work outlines a valuable protocol to study the binding of ligands to pLGICs beyond conventional docking and molecular dynamics techniques.