Yakovlev Gregory, Reda Torsten, Hirst Judy
Medical Research Council Dunn Human Nutrition Unit, Wellcome Trust/MRC Building, Hills Road, Cambridge CB2 2XY, United Kingdom.
Proc Natl Acad Sci U S A. 2007 Jul 31;104(31):12720-5. doi: 10.1073/pnas.0705593104. Epub 2007 Jul 19.
quinone oxidoreductase (complex I) plays a pivotal role in cellular energy production. It employs a series of redox cofactors to couple electron transfer to the generation of a proton-motive force across the inner mitochondrial or bacterial cytoplasmic membrane. Complex I contains a noncovalently bound flavin mononucleotide at the active site for NADH oxidation and eight or nine iron-sulfur clusters to transfer electrons between the flavin and a quinone-binding site. Understanding the mechanism of complex I requires the properties of these clusters to be defined, both individually and as an ensemble. Most functional information on the clusters has been gained from EPR spectroscopy, but some clusters are not observed by EPR and attributing the observed signals to the structurally defined clusters is difficult. The current consensus picture relies on correlating the spectra from overexpressed subunits (containing one to four clusters) with those from intact complexes I. Here, we analyze spectra from the overexpressed NuoG subunit from Escherichia coli complex I and compare them with spectra from the intact enzyme. Consequently, we propose that EPR signals N4 and N5 have been misassigned: signal N4 is from NuoI (not NuoG) and signal N5 is from the conserved cysteine-ligated [4Fe-4S] cluster in NuoG (not from the cluster with a histidine ligand). The consequences of reassigning the EPR signals and their associated functional information on the free energy profile for electron transfer through complex I are discussed.
醌氧化还原酶（复合体I）在细胞能量产生中起关键作用。它利用一系列氧化还原辅助因子将电子传递与线粒体内膜或细菌细胞质膜上质子动力的产生相偶联。复合体I在其活性位点含有一个非共价结合的黄素单核苷酸用于NADH氧化，以及八个或九个铁硫簇，用于在黄素和醌结合位点之间传递电子。要理解复合体I的机制，需要分别并整体地确定这些簇的性质。关于这些簇的大多数功能信息是通过电子顺磁共振光谱法获得的，但有些簇无法通过电子顺磁共振观察到，而且将观察到的信号归因于结构明确的簇很困难。目前的共识图景依赖于将过表达亚基（含有一到四个簇）的光谱与完整复合体I的光谱进行关联。在这里，我们分析了来自大肠杆菌复合体I过表达的NuoG亚基的光谱，并将其与完整酶的光谱进行比较。因此，我们提出电子顺磁共振信号N4和N5被错误归属：信号N4来自NuoI（而非NuoG），信号N5来自NuoG中保守的半胱氨酸连接的[4Fe - 4S]簇（而非来自具有组氨酸配体的簇）。讨论了重新归属电子顺磁共振信号及其相关功能信息对通过复合体I进行电子转移的自由能分布的影响。
