DISTANCE MEASUREMENTS IN METALLOPROTEINS

金属蛋白质的距离测量

• 批准号: 2178369
• 负责人: GARY W BRUDVIG
• 金额: $ 16.46万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-09-23 至 1995-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2178369
• 关键词: active sites; bacterial proteins; bioenergetics; biological transport; chelating agents; chlorophyll; copper; cytochrome b; cytochrome oxidase; electron spin resonance spectroscopy; electron transport; iron; magnetism; manganese; membrane proteins; metal complex; metalloproteins; molecular dynamics; oxidation reduction reaction; photosynthetic reaction centers; protein structure function; tyrosine

Energy transduction in respiration and photosynthesis involves the coupling of electron transfer reactions to the generation of an electrochemical gradient across a bilayer membrane. Although the sequence and kinetics of the electron transfer reactions are by and large known in these systems, much less is known about the spatial arrangement of the electron carriers and the distances between them. Ideally, the electron carriers could be used as probes in order to unravel their spatial organization and the distances of long-range electron transfer in the biological electron-transport processes. The long-term goals of this project are to use electron spin relaxation enhancement measurements to determine distances between redox active sites in proteins and to probe the magnetic properties of multinuclear metal ion clusters. The first objective is to characterize the effects of pairwise magnetic interactions on the electron spin relaxation rates in samples in which the distance between interacting centers and the magnetic properties of each center are known. In this study, the reaction center protein from Rb sphaeroides will be used as a model system. This protein can be prepared in a state that contains two paramagnetic sites at a fixed distance of 28 Angstroms. The electron spin relaxation rate enhancement of the bacteriochlorophyll special pair cation radical due to a magnetic interaction with the nonheme Fe, or Mn2+ or CU2+ in the Fe -site, will be systematically investigated by varying the metal ions and the temperature in both non-oriented and oriented samples. These data will be used to test and extend the theories for spin relaxation enhancement. The second objective is to use measurements of electron spin-lattice relaxation rate enhancements to determine the spatial organization and magnetic properties of redox sites in cytochrome c oxidase and photosystem II, two integral membrane protein complexes for which structural information is not available. Three types of applications will be pursued in order to obtain structural information in these two systems and also to establish the general applicability of this method to determine distances in other systems. The first application is to measure magnetic interactions between endogenous sites. The second is to measure interactions between endogenous sites and an exogenous Dy3+-chelate complex. The third is to use measurements of the temperature dependence of the spin-lattice relaxation rate of the stable tyrosine radical in Photosystem ii to probe the magnetic properties of the Mn cluster. These experiments will provide a basis for using electron spin-lattice relaxation measurements to determine distances of electron transfer and magnetic properties of metal ion clusters in other redox protein complexes.

呼吸和光合作用的能量转导涉及 电子转移反应与生成的耦合 跨双层膜的电化学梯度。 虽然 电子传递反应的序列和动力学是很大程度上的 在这些系统中已知，关于空间布置知之甚少 电子载体和它们之间的距离。 理想情况下， 电子载体可以用作探针以揭开其 空间组织和远程电子转移的距离 生物电子传输过程。 这个长期目标 项目将使用电子自旋松弛增强测量值 确定蛋白质中的氧化还原活性位点与探针之间的距离 多核金属离子簇的磁性。 第一个 目的是表征成对磁的效果 样品中电子自旋松弛率的相互作用 相互作用中心与磁性特性之间的距离 每个中心都是已知的。 在这项研究中，反应中心蛋白来自 RB Sphaeroides将用作模型系统。 该蛋白可以是 在包含两个固定的顺磁性位点的状态下制备 28埃埃斯特群的距离。 电子自旋松弛率提高 由于磁性而导致的细菌氯叶藻特殊对阳离子的自由基 与非血红素Fe的相互作用，或者在Fe-点中的MN2+或Cu2+相互作用，将是 通过改变金属离子和温度来系统地研究 在非方向和定向样品中。 这些数据将用于 测试并扩展理论以增强自旋松弛。 第二个 目的是使用电子自旋晶格松弛率的测量 确定空间组织和磁性的增强功能 细胞色素C氧化酶和光系统II中的氧化还原位点的特性，两个 结构信息的整体膜蛋白复合物 无法使用。 为了进行三种类型的申请 在这两个系统中获取结构信息，也建立 该方法确定其他距离的一般适用性 系统。 第一个应用是测量磁相互作用 内源地点之间。 第二个是测量 内源部位和外源性DY3+氯酸盐复合物。 第三是 使用旋转时间的温度依赖性的测量 稳定酪氨酸自由基在光系统II中的放松率 MN簇的磁性特性。 这些实验会 提供了使用电子自旋晶格松弛测量的基础 确定电子传输的距离和磁性 其他氧化还原蛋白复合物中的金属离子簇。