Projection structure of the membrane domain of Escherichia coli respiratory complex I at 8Å resolution
Projection structure of the membrane domain of Escherichia coli respiratory complex I at 8Å resolution
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DOI：
10.1016/j.jmb.2006.11.026
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发表时间：
2007-02-09
影响因子：
5.6
通讯作者：
Sazanov, Leonid A.
中科院分区：
文献类型：
--
作者：
Baranova, Ekaterina A.;Holt, Peter J.;Sazanov, Leonid A.
呼吸复合物 I（NADH：泛醌氧化还原酶）是一种 L 形多亚基蛋白质组装体，由疏水性膜臂和亲水性外周臂组成。它催化两个电子从 NADH 到醌的转移，以及四个质子的易位。穿过膜。虽然我们最近已经解决了外周臂的晶体结构，但完整酶的结构和偶联机制尚不清楚。大肠杆菌复合体 I 的膜结构域由 7 个不同的亚基组成，总分子质量为 258 kDa。它比整个酶更稳定，这使我们能够获得属于空间群 p22(1)2(1) 的结构域的有序二维晶体。将负染色晶体的投影图与先前发表的低分辨率结构进行比较表明，膜域的特征弯曲形状在细菌和线粒体酶之间非常保守，这有助于我们在完整复合物的背景下解释投影图。膜域远端的两个明显的染色排除密度可能代表两个大的逆向转运蛋白样亚基 NuoL 和 NuoM。冷冻水合晶体上的冷冻电子显微镜使我们能够计算 8 A 分辨率的投影图。一个膜域内存在大约 60 个垂直于膜平面和倾斜的跨膜 α 螺旋，这与二级结构预测一致。在与外围臂的界面处发现了醌的可能结合位点和进入通道。已经将各个子单元初步分配给地图的特征。亚基 NuoL 和 NuoM 的位置与外臂（包含复合物的所有氧化还原中心）相距相当远，表明构象变化可能在电子转移和质子泵浦之间的耦合机制中发挥作用。 (c) 2006 Elsevier Ltd. 保留所有权利。
Respiratory complex I (NADH:ubiquinone oxidoreductase) is an L-shaped multisubunit protein assembly consisting of a hydrophobic membrane arm and a hydrophilic peripheral arm. It catalyses the transfer of two electrons from NADH to quinone coupled to the translocation of four protons. across the membrane. Although we have solved recently the crystal structure of the peripheral arm, the structure of the complete enzyme and the coupling mechanism are not yet known. The membrane domain of Escherichia coli complex I consists of seven different subunits with total molecular mass of 258 kDa. It is significantly more stable than the whole enzyme, which allowed us to obtain well-ordered two-dimensional crystals of the domain, belonging to the space group p22(1)2(1). Comparison of the projection map of negatively stained crystals with previously published low-resolution structures indicated that the characteristic curved shape of the membrane domain is remarkably well conserved between bacterial and mitochondrial enzymes, helping us to interpret projection maps in the context of the intact complex. Two pronounced stain-excluding densities at the distal end of the membrane domain are likely to represent the two large antiporter-like subunits NuoL and NuoM. Cryo-electron microscopy on frozen-hydrated crystals allowed us to calculate a projection map at 8 A resolution. About 60 transmembrane alpha-helices, both perpendicular to the membrane plane and tilted, are present within one membrane domain, which is consistent with secondary structure predictions. A possible binding site and access channel for quinone are found at the interface with the peripheral arm. Tentative assignment of individual subunits to the features of the map has been made. The location of subunits NuoL and NuoM at substantial distance from the peripheral arm, which contains all the redox centres of the complex, indicates that conformational changes are likely to play a role in the mechanism of coupling between electron transfer and proton pumping. (c) 2006 Elsevier Ltd. All rights reserved.