Pohl Thomas, Bauer Theresa, Dörner Katerina, Stolpe Stefan, Sell Philipp, Zocher Georg, Friedrich Thorsten
Institut für Organische Chemie und Biochemie, Albert-Ludwigs-Universität Freiburg, Albertstrasse 21, Chemiehochhaus, D-79104 Freiburg i. Br., Germany.
Biochemistry. 2007 Jun 5;46(22):6588-96. doi: 10.1021/bi700371c. Epub 2007 May 10.
The NADH:ubiquinone oxidoreductase (complex I) from Escherichia coli is composed of 13 subunits called NuoA through NuoN. It catalyzes the electron transfer from NADH to ubiquinone by a chain of redox groups consisting of one FMN and seven iron-sulfur clusters. The function of the additional, nonconserved cluster N7 located on NuoG is not known. It has been speculated that it is not involved in electron transfer, due to its distance of more than 20 A from the electron transfer chain. Dithionite-reduced minus NADH-reduced EPR difference spectra of complex I and of a soluble fragment containing NuoG revealed for the first time the EPR spectrum of N7 in the complex. Individual mutation of the cysteines ligating this cluster to alanine led to a decreased amount of complex I in the membrane without affecting the electron transfer activity. Sucrose gradient centrifugation revealed that the complex from the C230A and C233A mutants decayed in detergent solution while the C237A and C265A mutant complex was stable. Cluster N7 was detectable in the latter mutants but with shifted g-values, indicating a different ligation of N7. Thus, N7 is essential for the stability of the complex but is not involved in electron transfer.
来自大肠杆菌的NADH：泛醌氧化还原酶（复合体I）由13个亚基组成，分别称为NuoA至NuoN。它通过由一个FMN和七个铁硫簇组成的氧化还原基团链催化电子从NADH转移到泛醌。位于NuoG上的额外的、非保守簇N7的功能尚不清楚。据推测，由于它与电子传递链的距离超过20埃，它不参与电子传递。复合体I以及包含NuoG的可溶性片段的连二亚硫酸盐还原减去NADH还原的电子顺磁共振（EPR）差异光谱首次揭示了该复合体中N7的EPR光谱。将连接该簇的半胱氨酸逐个突变为丙氨酸导致膜中复合体I的量减少，而不影响电子传递活性。蔗糖梯度离心显示，来自C230A和C233A突变体的复合体在去污剂溶液中降解，而C237A和C265A突变体复合体是稳定的。在后者的突变体中可检测到簇N7，但g值发生了偏移，表明N7的连接方式不同。因此，N7对复合体的稳定性至关重要，但不参与电子传递。
