RUI: The Cytochrome bf complex: Role in Redox Regulation and Function of the Quinone-reductase Site

RUI：细胞色素 bf 复合物：氧化还原调节中的作用和醌还原酶位点的功能

• 批准号: 0450875
• 负责人: Toivo Kallas
• 金额: --
• 依托单位: University of Wisconsin-Oshkosh
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-06-01 至 2010-05-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0450875&HistoricalAwards=false
• 关键词: RUI; Cytochrome; bf; complex; Role

The cytochrome bf complex of cyanobacterial, algal, and plant oxygenic photosynthesis transfers electrons between the two photochemical reaction centers and establishes a proton gradient for ATP synthesis. In addition, the bf complex plays a crucial role in redox sensing and regulation of photosynthesis. The catalytic mechanism involves plastoquinol oxidation on one side of the membrane and plastoquinone reduction on the other side. Quinone-reductase-site function is poorly understood but seems necessary for turnover of the bf complex, cyclic electron flow, and has implications for oxygen radical production. A unique, recently discovered heme (ci or x) resides in the quinone-reductase pocket. This project will investigate the mechanism and consequences of redox regulation by the bf complex and quinone-reductase-site function. Most studies will be performed using the cyanobacterium Synechocystis PCC 6803, which is amenable to genetics, time-resolved spectroscopy, and microarray analysis. Specific objectives are: 1) To investigate available Rieske iron-sulfur protein and low and high-potential chain mutants of the bf complex for state-transitions (the redistribution of light harvesting antenna) and electron transport kinetics. 2) To investigate specific mutants by microarray hybridizations, 2D gel separations, and mass spectrometry to characterize the role of the bf complex in gene and protein regulation. 3) To obtain additional redox-regulation mutants by chemical mutagenesis and fluorescence screening. 4) To construct structure-based, site-directed mutations in the cytochrome bf quinonereductase domain to investigate its catalytic mechanism and role in cyclic electron transport. Mutant cyanobacteria will be characterized for electron transfer kinetics, regulation of state-transitions, oxygen radical production, and properties of the ci (x) heme. The research is timely because of the cytochrome bf structures and will provide important new insights into protein structure-function and redox signaling in photosynthesis.Broader impacts: Research is an important component of the educational experience for many students at UW Oshkosh. The research on redox regulation, catalytic mechanisms, and oxygen radical production in the cytochrome bf complex of cyanobacterial photosynthesis is multifaceted, takes full advantage of the NSF-funded Proteomics and Functional-Genomics Core Facility at UW Oshkosh, and will provide numerous, excellent and exciting research opportunities for students in all aspects of the project.

蓝藻、藻类和植物氧光合作用的细胞色素bf复合物在两个光化学反应中心之间传递电子，并为ATP合成建立质子梯度。此外，bf复合物在光合作用的氧化还原传感和调控中起着至关重要的作用。催化机理包括膜一侧的质体醌氧化和另一侧的质体醌还原。醌-还原酶位点的功能尚不清楚，但似乎是bf络合物周转、循环电子流和氧自由基产生所必需的。一个独特的，最近发现的血红素（ci或x）居住在醌还原酶口袋。本项目将研究bf复合物和醌还原酶位点功能调控氧化还原的机制和后果。大多数研究将使用蓝细菌聚胞菌PCC 6803进行，这是适用于遗传学，时间分辨光谱和微阵列分析。具体目标是：1)研究可用的Rieske铁硫蛋白和bf复合物的低电位和高电位链突变体的状态转变（光收集天线的重新分配）和电子传输动力学。2)通过微阵列杂交、2D凝胶分离和质谱分析研究特异性突变体，以表征bf复合物在基因和蛋白质调控中的作用。3)通过化学诱变和荧光筛选获得更多的氧化还原调控突变体。4)构建基于结构的、位点导向的细胞色素bf醌化酶结构域突变，研究其在循环电子传递中的催化机制和作用。突变的蓝藻将被表征为电子转移动力学，状态转变的调节，氧自由基的产生，和ci (x)血红素的性质。由于细胞色素bf的结构，这项研究是及时的，将为光合作用中蛋白质的结构功能和氧化还原信号提供重要的新见解。更广泛的影响：研究是威斯康星大学奥什科什分校许多学生教育经历的重要组成部分。蓝藻光合作用细胞色素bf复合体的氧化还原调控、催化机制和氧自由基产生的研究是多方面的，充分利用了美国国家科学基金会资助的蛋白质组学和功能基因组学核心设施，并将在项目的各个方面为学生提供大量优秀和令人兴奋的研究机会。