Structure, biogenesis, and function of mitochondrial complex I

线粒体复合物 I 的结构、生物发生和功能

• 批准号: 283326566
• 负责人: Professor Dr. Volker Zickermann
• 金额: --
• 依托单位: Arbeitsgruppe Strukturelle Bioenergetik
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2015
• 资助国家: 德国
• 起止时间: 2014-12-31 至 2022-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/283326566?language=en
• 关键词: Structure; biogenesis; function; mitochondrial; complex

NADH:ubiquinone oxidoreductase (respiratory complex I) is a very large membrane protein complex with a central function in aerobic energy metabolism. The enzyme is known to release deleterious oxygen radicals under specific conditions causing e.g. tissue damage after myocardial infarction. Dysfunction of mitochondrial complex I is implicated in a number of neuromuscular diseases and neurodegenerative conditions. We have recently solved the X-ray structure of mitochondrial complex I from the aerobic yeast Yarrowia lipolytica at 3.6 to 3.9 Å resolution. The structure offered new and exciting insights into complex I function explaining energy conversion and long range energy transfer in the large enzyme complex. For a comprehensive understanding of redox-linked proton translocation higher resolution data are needed. We propose different strategies to improve crystallization and to generate alternative crystal forms by using antibody fragments or T4 lysozyme fusions of accessory subunits. The most problematic part of the current X-ray structure is located in the upper half of the hydrophilic matrix arm of complex I. We have developed a scheme for controlled separation and purification of the soluble peripheral arm and propose to crystallize it as a discrete entity.We hypothesized that concerted structural changes at the ubiquinone reduction site trigger proton translocation. We propose to crystallize complex I variants carrying mutations of key residues to obtain insight into the structural basis of redox-linked proton translocation. The accessory NUEM subunit of complex I belongs to the large family of short-chain dehydrogenases and contains an NADPH binding site. We aim to unravel the yet unknown function of NUEM in mitochondrial metabolism and to resolve the possible interaction of NUEM with a complex I bound acyl-carrier protein.Complex I biogenesis is a complex multi-step process. Recently, we have isolated and studied an assembly intermediate that binds assembly factor N7BML (human NDUFAF2) and we hypothesized that N7BML is needed to allow interaction with the iron-sulfur cluster insertion machinery during assembly. We propose to determine the structure of the assembly intermediate by high-resolution electron microscopy.

NADH：泛醌氧化还原酶(呼吸复合体I)是一种非常大的膜蛋白复合体，在有氧能量代谢中起中心作用。众所周知，这种酶在特定条件下会释放有害的氧自由基，导致心肌梗死后的组织损伤。线粒体复合体I功能障碍与许多神经肌肉疾病和神经退行性疾病有关。我们最近解开了解脂雅罗维菌需氧酵母线粒体复合体I的X-射线结构，分辨率为3.6到3.9。该结构为研究复杂的I功能提供了新的和令人兴奋的见解，解释了大型酶复合体中的能量转换和远程能量转移。为了全面了解氧化还原相关的质子转移，需要更高分辨率的数据。我们提出了不同的策略来改善结晶，并通过使用抗体片段或辅助亚基的T4溶菌酶融合来产生不同的晶体形式。目前X射线结构中最有问题的部分位于配合物I的亲水基质臂的上半部分。我们开发了一种控制分离和纯化可溶外周臂的方案，并建议将其结晶为一个离散的实体。我们假设泛醌还原位置的协同结构变化触发了质子转移。我们建议将携带关键残基突变的复合体I变体结晶，以深入了解氧化还原连接的质子转运的结构基础。复合体I的辅助NUEM亚基属于短链脱氢酶大家族，含有NADPH结合位点。我们的目标是揭开NUEM在线粒体代谢中的未知功能，并解决NUEM可能与复杂的I结合的酰基载体蛋白相互作用的问题。最近，我们分离并研究了一种与组装因子N7BML(人NDUFAF2)结合的组装中间体，我们假设在组装过程中需要N7BML来允许与铁-硫簇插入机械的相互作用。我们建议用高分辨电子显微镜来确定组装中间体的结构。