Putting Molecular Dynamics to the Test: Ion Permeation

描述（由申请人提供）：本项目的目标是评估计算方法，通过紧密结合实验（电生理学、热力学和NMR）信息和基于原子模型（全原子分子动力学，MD;平均力的多离子势，PMF;布朗动力学，BD）分层实施的计算，深入了解离子渗透的机理。为了充分利用理论和实验之间的协同作用，这样的调查，重点必须放在一个特别明确的离子通道。我们选择短杆菌肽通道是因为短杆菌肽通道足够小，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-并且它们在某种意义上是不平凡的，因为它们的渗透性特性已被证明难以定量模拟。短杆菌肽通道，从而成为系统的选择，探索是否有可能发展一个全面的理解，在分子水平上使用原子计算的离子渗透。 拟议的研究是三个实验室之间的合作努力，它们为项目提供了补充专门知识，并建立了富有成效的合作记录。最初的计算研究将使用一个大的现有基础的测量单通道电流作为渗透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。平行地，短杆菌肽类似物将被合成以在PMF中引入限定的扰动，用于离子移动通过通道：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。取代的结构结果将使用固态NMR来确定。将使用单通道测量确定功能后果。这些类似物将进行MD和BD模拟-通道/侧链结构和动力学模拟，其中NMR结果将作为参考;以及离子渗透模拟，其中电生理学结果将作为参考。将测量碱金属阳离子从水到N-甲基乙酰胺的转移自由能，并用于确定离子-肽基团相互作用的适当表示，并帮助校准/改善原子力场。拟议的研究将有助于建立目前的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.