Enzyme design and protein dynamics.

在这个程序项目中以前的工作已经确定了参与催化功能的运动， 内切酶我们在这一更新应用中的工作假设是，通过使用理论和 计算方法，我们可以确定结构元素，创造动力学相关的催化作用， 所有的时间尺度，通过这些手段，我们可以创建蛋白质动力学设计范式， 增强和酶抑制。为了完成这项研究计划，我们将在5个不同的工作 这些领域是：1）我们将确定反应在所有时间尺度上所需的构象运动。正如 我们已经跟踪了轨迹，并确定了形成促进振动的蛋白质动力学， 反应坐标，我们将这些计算扩展到酶周转的时间尺度。2)我们将 鉴定特定酶系统中结构元件（乳酸脱氢酶和嘌呤核苷 磷酸化酶），产生特定的动力学。3)我们将确定变构结合位点，并确定 结合对蛋白质动力学的影响。4)我们将确定动力学如何参与过渡态 抑制剂结合。5)我们将建议重新设计“重型”PNP，以纠正我们现有的动态缺陷， 以前确定的和恢复正常的催化功能。 在这个计划项目中，所有工作的总推力是将动力学识别为 酶功能的中心特征，从选择催化活性构象底物， 快速促进振动。我们现在进入该计划的下一步-将动力学识别为 酶功能的设计元素。我们建议开始将这一概念发展为蛋白质的过程 工程工具

Previous work in this program project has identified motions that are involved in the catalytic function of enzymes. The hypothesis for our work in this renewal application is that by using theoretical and computational methods we can identify the structural elements that create dynamics related to catalysis on all timescales, and through these means we may create protein dynamics design paradigms for both rate enhancement, and enzyme inhibition. In order to complete this research program we will work on 5 distinct areas that are: 1) we will identify conformational movements necessary on all timescales for reaction. Just as we have followed trajectories and identified protein dynamics that form promoting vibrations and are part of the reaction coordinate, we will extend these calculations to the timescale of enzyme turnover. 2) We will identify structural elements in specific enzyme systems (lactate dehydrogenase and purine nucleoside phosphorylase) that create the specific dynamics. 3) We will identify allosteric binding sites and determine the effect of binding on protein dynamics. 4) We will identify how dynamics is involved in transition state inhibitor binding. 5) we will propose a redesign of "heavy" PNP that corrects dynamic defects we have previously identified and restores normal catalytic function. The overall thrust to all of the work in this program project has been the identification of dynamics as a central feature in enzyme function from the selection of a catalytically competent conformational substate to rapid promoting vibrations. We now proceed to the next step in this program - identification of dynamics as a design element in enzyme function. We propose to begin the process of developing this concept as a protein engineering tool.