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

RUI：金属蛋白短程电子转移的分子机制

• 批准号: 1817448
• 负责人: Oleksandr Kokhan
• 金额: $ 44.99万
• 依托单位: James Madison University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-07-01 至 2022-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1817448&HistoricalAwards=false
• 关键词: RUI; Molecular; Mechanisms; Short; Range

This project will focus 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 the 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 will support innovations in the science curriculum.The system that will be studied is PpcA, a 3 heme cytochrome from Geobacter sulfurreducens, genetically modified and covalently labeled with a number of photosensitizers will be used as a model system to study charge transfer reactions. 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 a number of structural techniques. 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 will also facilitate curricular updates in a number of upper level laboratory courses at James Madison University.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.

本项目将集中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，该项目还将为本科生提供研究培训，并将支持科学课程的创新。将研究的系统是PPCA，一种来自硫还原地杆菌的3-血红素细胞色素，经过转基因并共价标记了一些光敏剂，将作为研究电荷转移反应的模型系统。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。这些预测将通过一些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。该项目还将促进詹姆斯·麦迪逊大学一些高级实验室课程的课程更新。该奖项反映了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。

项目成果

Detection and quantification of transition metal leaching in metal affinity chromatography with hydroxynaphthol blue

• DOI: 10.1016/j.ab.2019.113347
• 发表时间: 2019-10-01
• 期刊: ANALYTICAL BIOCHEMISTRY
• 影响因子: 2.9
• 作者: Kokhan, Oleksandr;Marzolf, Daniel R.
• 通讯作者: Marzolf, Daniel R.

Examination of abiotic cofactor assembly in photosynthetic biomimetics: site-specific stereoselectivity in the conjugation of a ruthenium(II) tris(bipyridine) photosensitizer to a multi-heme protein

光合仿生学中非生物辅因子组装的检查：钌(II)三联吡啶光敏剂与多血红素蛋白缀合的位点特异性立体选择性

• DOI: 10.1007/s11120-019-00697-8
• 发表时间: 2020
• 期刊: Photosynthesis Research
• 影响因子: 3.7
• 作者: Ponomarenko, Nina S.;Kokhan, Oleksandr;Pokkuluri, Phani R.;Mulfort, Karen L.;Tiede, David M.
• 通讯作者: Tiede, David M.

Quantification of Metal Leaching in Immobilized Metal Affinity Chromatography

固定化金属亲和色谱法中金属浸出的定量

• DOI: 10.3791/60690
• 发表时间: 2020
• 期刊: Journal of Visualized Experiments
• 作者: Swaim, Coleman M.;Brittain, Tyler J.;Marzolf, Daniel R.;Kokhan, Oleksandr
• 通讯作者: Kokhan, Oleksandr