Ngo Van, Wang Yibo, Haas Stephan, Noskov Sergei Y, Farley Robert A
Department of Physics and Astronomy, University of Southern California, Los Angeles, California, United States of America.
Department of Biological Sciences, Centre for Molecular Simulations, University of Calgary, Calgary, Alberta, Canada.
PLoS Comput Biol. 2016 Jan 4;12(1):e1004482. doi: 10.1371/journal.pcbi.1004482. eCollection 2016 Jan.
Crystal structures of several bacterial Na(v) channels have been recently published and molecular dynamics simulations of ion permeation through these channels are consistent with many electrophysiological properties of eukaryotic channels. Bacterial Na(v) channels have been characterized as functionally asymmetric, and the mechanism of this asymmetry has not been clearly understood. To address this question, we combined non-equilibrium simulation data with two-dimensional equilibrium unperturbed landscapes generated by umbrella sampling and Weighted Histogram Analysis Methods for multiple ions traversing the selectivity filter of bacterial Na(v)Ab channel. This approach provided new insight into the mechanism of selective ion permeation in bacterial Na(v) channels. The non-equilibrium simulations indicate that two or three extracellular K+ ions can block the entrance to the selectivity filter of Na(v)Ab in the presence of applied forces in the inward direction, but not in the outward direction. The block state occurs in an unstable local minimum of the equilibrium unperturbed free-energy landscape of two K+ ions that can be 'locked' in place by modest applied forces. In contrast to K+, three Na+ ions move favorably through the selectivity filter together as a unit in a loose "knock-on" mechanism of permeation in both inward and outward directions, and there is no similar local minimum in the two-dimensional free-energy landscape of two Na+ ions for a block state. The useful work predicted by the non-equilibrium simulations that is required to break the K+ block is equivalent to large applied potentials experimentally measured for two bacterial Na(v) channels to induce inward currents of K+ ions. These results illustrate how inclusion of non-equilibrium factors in the simulations can provide detailed information about mechanisms of ion selectivity that is missing from mechanisms derived from either crystal structures or equilibrium unperturbed free-energy landscapes.
最近已发表了几种细菌钠通道的晶体结构，并且通过这些通道进行离子渗透的分子动力学模拟与真核通道的许多电生理特性一致。细菌钠通道的功能被表征为不对称，而这种不对称的机制尚未得到清楚理解。为了解决这个问题，我们将非平衡模拟数据与通过伞形采样和加权直方图分析方法生成的二维平衡无扰态势能面相结合，用于多个离子穿过细菌钠通道Na(v)Ab的选择性过滤器。这种方法为细菌钠通道中选择性离子渗透的机制提供了新的见解。非平衡模拟表明，在向内方向施加力的情况下，两到三个细胞外钾离子可以阻断Na(v)Ab选择性过滤器的入口，但在向外方向则不会。阻断状态出现在两个钾离子平衡无扰自由能态势能面的不稳定局部最小值处，适度的外力可以将其“锁定”在该位置。与钾离子相反，三个钠离子以松散的“连锁”渗透机制作为一个单元在向内和向外方向上顺利通过选择性过滤器，并且在两个钠离子的二维自由能态势能面中不存在类似的阻断状态局部最小值。非平衡模拟预测的打破钾离子阻断所需的有用功相当于实验测量的两种细菌钠通道诱导钾离子内向电流所需的大外加电位。这些结果说明了在模拟中纳入非平衡因素如何能够提供有关离子选择性机制的详细信息，而这些信息是从晶体结构或平衡无扰自由能态势能面推导的机制中所缺失的。
