Coupling between conformation and chemistry in enzymes

描述（由申请人提供）：肌球蛋白是原型分子马达的超家族，在从囊泡运输、细胞运动到肌肉收缩和信号转导的各种生物过程中发挥重要作用。虽然肌球蛋白的功能周期是了解在一个轮廓的形式，许多详细的问题仍然是关于运动域的构象特性和ATP酶活性之间的耦合。 我们推测，肌球蛋白门ATP水解的构象转变不仅通过调节活性位点中特定氨基酸的位置，而且通过调节水解位点周围水分子的方向和动力学。为了验证和巩固这样一个假设，国家的最先进的分子模拟提出了分析ATP水解的机制在不同的构象状态的肌球蛋白II马达结构域和相关的突变体，模拟包括经典的分子动力学和结合QM/MM方法。具体目的是：（1）确定运动域封闭状态下ATP水解的催化机制。(2)确定ATP水解是否在运动域的开放状态下被禁止，如果是，确定决定水解能量学的开放和闭合构象之间的关键差异。(3)用能量和机械的术语解释活性位点残基的作用，这些残基已被诱变研究证明对ATP水解和运动有各种影响。 选择肌球蛋白-II是因为它是唯一的运动系统，具有高分辨率结构的多个构象状态，计算结果可以与大量的生物化学和生物物理数据进行比较。拟议的模拟研究将提供一个框架，桥接来自不同学科的实验数据，建立合理的理论模型，在肌球蛋白和其他分子马达的机械化学耦合;微观的见解将对我们的能力产生深远的影响，设计策略，以治疗肌球蛋白功能障碍，如心肌病引起的严重疾病。 模拟工作将通过合作与实验研究紧密结合;结构，动力学和运动数据的组合将提供必要的实验测试来验证和改进模拟技术，这对计算酶学领域具有巨大价值。

DESCRIPTION (provided by applicant): Myosin is a superfamily of prototypical molecular motors that play important roles in diverse biological processes ranging from vesicle trafficking, cell motility to muscle contractions and signal transductions. Although the functional cycle of myosins is understood in an out-line form, many detailed questions remain concerning the coupling between conformational properties of the motor domain and the ATPase activity. We hypothesize that conformational transitions in myosin gate ATP hydrolysis through regulating not only positions of specific amino acids in the active site but also the orientation and dynamics of water molecules surrounding the hydrolysis site. To verify and consolidate such a hypothesis, state-of-the-art molecular simulations are proposed to analyze the mechanism of ATP hydrolysis in different conformational states of the myosin II motor domain and relevant mutants; the simulations include classical molecular dynamics and combined QM/MM methods. The specific aims are: (1) Determine the catalytic mechanism of ATP hydrolysis in the closed state of the motor domain. (2) Determine if ATP hydrolysis is prohibited in the open state of the motor domain, and if so, identify key differences between the open and closed conformations that dictate the hydrolysis energetics. (3) Explain, in energetical and mechanistic terms, the roles of active site residues, which have been shown by mutagenesis studies to have various effects on ATP hydrolysis and motility. Myosin-ll was chosen because it is the only motor system that has high-resolution structures for multiple conformational states, and computational results can be compared with a large body of biochemical and biophysical data. The proposed simulation study will provide a framework for bridging experimental data from different disciplines to establish sensible theoretical models for mechanochemical coupling in myosin and other molecular motors; the microscopic insights will have a profound impact on our ability to design strategies to treating serious diseases caused by myosin dysfunction such as cardiomyopathy. The simulation work will be closely coupled to experimental studies through collaborations; the combination of structural, kinetic and motility data will provide the experimental tests necessary to verify and refine simulation techniques, which is of tremendous value to the field of computational enzymology.