Biophysical and structural characterization of proton-translocating NADH-dehydrogenase (complex I) from the strictly aerobic yeast Yarrowia lipolytica
Biophysical and structural characterization of proton-translocating NADH-dehydrogenase (complex I) from the strictly aerobic yeast Yarrowia lipolytica
复制标题
DOI：
10.1016/s0005-2728(00)00154-7
复制
发表时间：
2000-07-20
影响因子：
4.3
通讯作者：
Brandt, U
中科院分区：
文献类型：
--
作者：
Djafarzadeh, R;Kerscher, S;Brandt, U
线粒体质子转位 NADH 脱氢酶（复合物 I）是最大、最复杂的膜结合蛋白复合物之一。尽管它在真核氧化磷酸化中发挥核心作用并参与广泛的人类疾病，但对其结构和功能知之甚少。因此，我们开始使用可用于严格需氧酵母解脂耶氏酵母的强大遗传工具来研究这种呼吸链酶。为了建立解脂耶氏酵母作为复合物 I 的模型系统，我们纯化并表征了解脂耶氏酵母的多亚基酶，并对编码其外周部分的七个中心亚基的核基因进行了测序。来自解脂耶氏酵母的复合物 I 非常稳定，可以以高纯度和单分散状态分离。 EPR 光谱检测到了 1 个双核和 4 个四核铁硫簇，包括以前仅从哺乳动物线粒体中发现的 N5。在负染色和冰中通过单颗粒电子显微镜进行的初步结构分析表明，来自解脂耶氏酵母的复合物 I 为 L 形颗粒，在外围和膜部分之间没有表现出在其他系统中观察到的细茎。 (C) 2000 Elsevier Science B.V. 保留所有权利。
Mitochondrial proton-translocating NADH-dehydrogenase (complex I) is one of the largest and most complicated membrane bound protein complexes. Despite its central role in eukaryotic oxidative phosphorylation and its involvement in a broad range of human disorders, little is known about its structure and function. Therefore, we have started to use the powerful genetic tools available for the strictly aerobic yeast Yarrowia lipolytica to study this respiratory chain enzyme. To establish Y. lipolytica as a model system for complex I, we purified and characterized the multisubunit enzyme from Y. lipolytica and sequenced the nuclear genes coding for the seven central subunits of its peripheral part. Complex I from Y. lipolytica is quite stable and could be isolated in a highly pure and monodisperse state. One binuclear and four tetranuclear iron-sulfur clusters, including N5, which was previously known only from mammalian mitochondria, were detected by EPR spectroscopy. Initial structural analysis by single particle electron microscopy in negative stain and ice shows complex I from Y. lipolytica as an L-shaped particle that does not exhibit a thin stalk between the peripheral and the membrane parts that has been observed in other systems. (C) 2000 Elsevier Science B.V. All rights reserved.