RUI: Molecular Mechanisms of Short-Range Electron Transfer in Metalloproteins

本项目的重点是了解蛋白质中短程电子转移的分子水平机制。所有的生命系统都是通过占据高能态的电子获得能量的，要么通过呼吸链（食物），要么通过光（光合作用）。电子转移是许多负责储存、转移和转化能量的蛋白质的重要功能。虽然对控制长距离电荷转移的因素有一些了解，但对短于1纳米的生物电子转移知之甚少。填补这一空白将推进我们对生物能量学基本步骤的认识。通过建立计算和实验模型来预测和解释这些反应的关键参数，将获得对短程生物电子转移的一个有凝聚力的理解。此外，该项目将为本科生和代表性不足的少数民族学生提供研究培训，并将支持科学课程的创新。该项目还将促进詹姆斯麦迪逊大学几门高级实验课程的课程更新。PpcA是一种来自硫还原地杆菌的3血红素c型细胞色素，经过基因修饰并与几种光敏剂共价标记，将被用作研究电荷转移反应的模型体系。总体假设是紧密的结构耦合和多余热能的有效耗散是实现超快电荷转移速率的必要条件。反应的动力学将在室温和低温下用时间分辨荧光和吸收光谱进行研究。蛋白质-光敏剂复合物的结构完整性将通过SAXS和CD光谱进行监测。广泛的全原子分子动力学模拟将进行预测结构和评估结构动力学。这些预测将通过包括核磁共振光谱和x射线晶体学在内的几种结构技术进行实验测试。收集到的动力学数据将在分子结构的背景下进行分析，并将用于测试和修改目前可用的预测电子转移速率和途径的计算和理论方法。该项目由分子和细胞生物科学部的分子生物物理集群资助。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

This project is focused on understanding the molecular level mechanism of short-range electron transfer in proteins. All living systems obtain energy through electrons occupying high energy states, either through respiratory chains (food) or from light (photosynthesis). Electron transfer is a vital function of the many proteins responsible for storage, transfer, and transformation of this energy. While there is some understanding of the factors controlling charge transfer over longer distances, little is known about biological electron transfer at distances shorter than 1 nm. Filling this gap will advance our knowledge of the fundamental steps in bioenergetics. By developing computational and experimental models to predict and explain the key parameters of these reactions, a cohesive understanding of short-range biological electron transfer will be obtained. In addition, the project will provide research training for undergraduates and underrepresented minority students and will also support innovations in the science curriculum. This project will also facilitate curricular updates in several upper-level laboratory courses at James Madison University. PpcA, a 3 heme c-type cytochrome from Geobacter sulfurreducens, genetically modified and covalently labeled with several photosensitizers will be used as a model system to study charge transfer reactions. The overarching hypothesis is that tight structural coupling and effective dissipation of excess heat energy are essential to achieve ultrafast charge transfer rates. The kinetics of the reactions will be studied with time-resolved fluorescence and absorbance spectroscopies at room and cryogenic temperatures. The structural integrity of protein-photosensitizer complexes will be monitored with SAXS and CD spectroscopy. Extensive all-atom molecular dynamics simulations will be performed to predict structures and to evaluate structural dynamics. These predictions will be tested experimentally with several structural techniques including nuclear magnetic resonance spectroscopy and x-ray crystallography. The collected kinetic data will be analyzed in the context of molecular structures and will be used to test and revise currently available computational and theoretical approaches for the prediction of electron transfer rates and pathways. This project is funded by the Molecular Biophysics Cluster in the Division of Molecular and Cellular Biosciences.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.