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

本项目致力于了解蛋白质中短程电子转移的分子水平机制。所有的生命系统都通过占据高能态的电子来获得能量，要么是通过呼吸链(食物)，要么是通过光(光合作用)。电子转移是许多蛋白质的重要功能，这些蛋白质负责存储、转移和转换这种能量。虽然对控制较长距离的电荷转移的因素有一些了解，但对距离小于1纳米的生物电子转移知之甚少。填补这一空白将促进我们对生物能量学基本步骤的了解。通过开发计算和实验模型来预测和解释这些反应的关键参数，将对短程生物电子转移有一个连贯的理解。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室课程的更新。PPCA是一种来自硫还原地杆菌的3-血红素c型细胞色素，经过基因修饰后与几种光敏剂共价标记，将作为研究电荷转移反应的模型体系。最重要的假设是，紧密的结构耦合和过剩热能的有效耗散是实现超快电荷转移速率的关键。在室温和低温下，将用时间分辨荧光和吸收光谱研究反应的动力学。蛋白质-光敏剂复合体的结构完整性将用SAXS和CD光谱进行监测。将进行广泛的全原子分子动力学模拟来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射线结晶学在内的几种结构技术进行实验验证。收集的动力学数据将在分子结构的背景下进行分析，并将用于测试和修订目前可用于预测电子转移速率和路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组资助。这一奖项反映了NSF的法定使命，并通过使用基金会的智力优势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.