DISTANCE MEASUREMENTS IN METALLOPROTEINS

在呼吸和光合作用中的能量转换涉及 将电子转移反应与生成 电化学梯度穿过双层膜。 虽然 电子转移反应的顺序和动力学基本上是 在这些系统中已知的，关于空间布置的知之甚少 以及它们之间的距离。 最好 电子载体可以用作探针，以解开它们的 空间组织和远程电子转移的距离 生物电子传递过程。 长期目标是 项目是使用电子自旋弛豫增强测量， 确定蛋白质中氧化还原活性位点之间的距离， 多核金属离子团簇的磁性。 第一 目的是表征成对磁效应 相互作用对样品中电子自旋弛豫速率的影响， 相互作用中心之间的距离和磁性能 每个中心都是已知的。 在这项研究中，反应中心蛋白从 Rb sphaeroides将被用作模型系统。 这种蛋白质可以 在含有两个顺磁性位点的状态下制备， 距离28安。 电子自旋弛豫速率增强 细菌叶绿素特殊对阳离子自由基由于磁性 与非血红素Fe或Fe -位点中的Mn 2+或Cu 2+的相互作用将是 通过改变金属离子和温度， 在非定向和定向样品中。 这些数据将用于 测试和扩展自旋弛豫增强理论。 第二 目的是利用电子自旋-晶格弛豫速率的测量 增强以确定空间组织和磁 细胞色素c氧化酶和光系统II中氧化还原位点的性质， 完整的膜蛋白复合物，其结构信息 不可用. 我们会处理三类申请， 获得这两个系统的结构信息，并建立 这种方法在确定其他地区距离方面的普遍适用性 系统. 第一个应用是测量磁相互作用 在内源性位点之间。 第二个是测量 内源性位点和外源性Dy 3 +-螯合复合物。 三是 使用自旋晶格的温度依赖性的测量 光系统ii中稳定酪氨酸自由基的弛豫速率 Mn团簇的磁性。 这些实验将 为使用电子自旋晶格弛豫测量提供了基础 来确定电子转移的距离和磁性， 其他氧化还原蛋白质复合物中的金属离子簇。

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.