Putting Molecular Dynamics to the Test: Ion Permeation

描述(申请人提供)：这个项目的目标是评估计算方法，通过紧密结合来自实验(电生理学、热力学和核磁共振)的信息和基于原子模型(全原子分子动力学，MD；多离子平均作用力势，PMF；和布朗动力学，BD)的计算，获得对离子渗透的深入机理洞察。为了充分利用理论和实验之间的协同作用，这种研究的重点必须放在一个特别明确的离子通道上。我们之所以选择Graicidin通道，是因为Graicidin通道足够小，可以进行详细的模拟，但又足够大，它们具有明确的结构和功能特性-而且它们的渗透性很难被定量模拟，因此它们不是平凡的。因此，Gramiidin通道成为探索是否有可能利用原子计算在分子水平上全面了解离子渗透的首选系统。 拟议的研究是三个实验室之间的合作努力，这三个实验室为该项目提供互补的专业知识，并建立了富有成效的合作记录。最初的计算研究将使用现有的大量单通道电流测量作为平均离子浓度和跨膜电压的函数，作为通道介导的离子运动的原子模拟的参考。同时，将合成gricidin类似物，以在PMF中为离子通过通道的运动引入定义的扰动：Trp-“Phe取代，其中极性Trp被非极性Phe取代；Ala-”Ser取代，其中非极性Ala被极性Ser取代。取代的结构后果将使用固体核磁共振来确定。功能后果将使用单通道测量来确定。这些类似物将接受MD和BD模拟-通道/侧链结构和动力学，其中核磁共振结果将作为参考；以及离子渗透，其中电生理结果将作为参考。将测量碱金属阳离子从水到N-甲基乙酰胺的转移自由能，并将其用于确定离子-肽基相互作用的适当表示，并帮助校准/改进原子力场。拟议的研究将有助于确定当前离子渗透模型的有效性范围，确定任何差异的物理基础，并实施解决方案，以解决遇到的任何困难。

DESCRIPTION (provided by applicant): The goal of this project is to evaluate computational methods for obtaining in-depth mechanistic insight into ion permeation by tightly combining information from experiments (electrophysiology, thermodynamics and NMR) and computations based on a hierarchical implementation of atomistic models (all-atom molecular dynamics, MD; multi-ion potential of mean force, PMF; and Brownian dynamics, BD). For such investigations to fully exploit the synergism between theory and experiment, the focus must be on a particularly well-defined ion channel. We selected the gramicidin channel because gramicidin channels are small enough that it is feasible to undertake detailed simulations, yet large enough that they have well-defined structural and functional properties - and they are non-trivial in the sense that their permeability properties have proven difficult to simulate quantitatively. Gramicidin channels thus become the system of choice for exploring whether it is possible to develop a comprehensive understanding of ion permeation at the molecular level using atomistic calculations. The proposed studies represent a cooperative effort among three laboratories, which provide complementary expertise to the project and have established records of productive collaborations. The initial computational studies will use a large existing base of measurements of single-channel currents as functions of permeant ion concentration and transmembrane voltage, as a reference for atomistic simulations of channel-mediated ion movement. In parallel, gramicidin analogues will be synthesized to introduce defined perturbations in the PMF for ion movement through the channels: Trp-"Phe substitutions, where the polar Trp is replaced by the nonpolar Phe; Ala-"Ser substitutions, where the non-polar Ala is replaced by the polar Ser. The structural consequences of the substitutions will be determined using solid-state NMR. The functional consequences will be determined using single-channel measurements. These analogues will be subject to MD and BD simulations - of the channel/side chain structure and dynamics, where the NMR results will serve as reference; and of ion permeation, where the electrophysiological results will serve as reference. The transfer free energy of alkali metal cations from water to N-methyl-acetamide will be measured and used to ascertain appropriate representation of ion-peptide group interactions and help calibrate/improve the atomic force field. The proposed research will help establish the range of validity of current models of ion permeation, identify the physical basis of any discrepancy, and implement solutions to resolve any difficulties that are encountered.