SUBSTRATE DOCKING IN CYTOCHROME C OXIDASE

细胞色素 C 氧化酶中的底物对接

• 批准号: 6271921
• 负责人: SHELAGH M FERGUSON-MILLER
• 金额: $ 11.58万
• 依托单位: MICHIGAN STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-06-01 至 1999-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6271921
• 关键词: Raman spectrometry; Rhodospirillales; X ray crystallography; active sites; chemical binding; chemical kinetics; chemical structure function; chimeric proteins; cytochrome c; cytochrome oxidase; electron spin resonance spectroscopy; electron transport; enzyme activity; enzyme structure; enzyme substrate; intermolecular interaction; ionic bond; mathematical model; protein binding; site directed mutagenesis; structural biology; ultracentrifugation; water

Project IV- Substrate Docking in Cytochrome c Oxidase (Ferguson-Miller, Kuhn, Garavito, Roberts) Cytochrome c oxidase is an intrinsic membrane protein whose complex energy transducing function involves electron transfer, reduction of oxygen to water and translocation of protons across the membrane. This proposal is aimed at determining the nature of the protein-protein interaction between cytochrome c and cytochrome c oxidase, which determines the rate and efficiency of electron delivery in the oxidase. Models will be developed for the docking sties for cytochrome c on CcOX by a powerful computational algorithm that uses protein electrostatic fields and van der Waals surfaces in a systematic orientation search of the intermolecular energies of the two proteins. The predictions of this model will be tested by mutation of Rhodobacter sphaeroides CcOX, comparison of the kinetics and binding of cytochrome c with the mutant and wild-type enzymes, and by crystallizing native and mutant forms of Rhodobacter CcOX and CcOX/cytochrome c complexes, to determine their structure by X-ray analysis. Homology modeling of the Rhodobacter sequence into the bovine oxidase coordinates (available to us from our collaborator, S. Yoshikawa) will permit computational analysis of the docking with a model closer to that of the bacterial enzyme. Conversely, creation of a chimeric bacterial enzyme with a mammalian coxII gene substituted for the Rhodobacter gene, will allow assays of binding to be done in a system that better corresponds to the mammalian oxidase structure and better matches the mammalian cytochrome c. Surface water at the oxidase/cytochrome c interface will also be analyzed to predict retained water molecules that may influence the chemistry of the interaction. Defining the role of electrostatics and other forces in this protein-protein interaction will increase our understanding of the mechanism and mechanics of electron transfer in the respiratory chain.

项目IV-细胞色素c氧化酶中的底物对接（Ferguson-Miller， 库恩、加拉维托、罗伯茨） 细胞色素c氧化酶是一种内在的膜蛋白， 转导功能包括电子转移，氧的还原， 水和质子穿过膜的移位。这项建议是 旨在确定蛋白质之间相互作用的性质， 细胞色素c和细胞色素c氧化酶，它决定了速率， 在氧化酶中的电子传递效率。将开发模型 通过一个强大的计算平台， 使用蛋白质静电场和货车范德华力的算法 表面的分子间能量的系统取向搜索 这两种蛋白质。该模型的预测将通过以下方式进行测试： 球形红细菌CcOX的诱变，动力学和 细胞色素c与突变体和野生型酶的结合，以及 使红细菌CcOX的天然和突变形式结晶， CcOX/细胞色素c复合物，通过X射线确定其结构 分析. Rhodobacter序列在牛中的同源性建模 氧化酶坐标（可从我们的合作者S. Yoshikawa） 将允许计算分析的对接模型更接近 即细菌酶。相反，嵌合细菌的产生 用哺乳动物coxII基因取代红细菌属基因的酶， 将允许在一个系统中进行结合测定， 对应于哺乳动物氧化酶结构，并且更好地匹配 哺乳动物细胞色素c氧化酶/细胞色素c处的地表水 界面也将被分析，以预测保留的水分子， 可能会影响相互作用的化学反应。定义的作用 蛋白质-蛋白质相互作用中的静电和其他力将 增加我们对电子的机制和力学的理解 在呼吸链中传递。