Chevallet M, Dupuis A, Lunardi J, van Belzen R, Albracht S P, Issartel J P
Laboratoire de BioEnergétique Cellulaire et Pathologique, DBMS, Commissariat à l'Energie Atomique, Grenoble, France.
Eur J Biochem. 1997 Dec 1;250(2):451-8. doi: 10.1111/j.1432-1033.1997.0451a.x.
The nuoI gene that encodes a ferredoxin-like subunit of the Rhodobacter capsulatus Complex I (a subunit equivalent to the bovine TYKY subunit) was mutated by homologous recombination. Both a nuoI-deleted mutant (delta nuoI mutant) and a point mutant in which Cys74 was replaced by a serine (C74S mutant) proved to be completely deficient in Complex I activity. These strains were unable to grow under anaerobic photosynthetic conditions. Their cytoplasmic membranes were also characterized by the absence of specific EPR signals assigned to FeS clusters N1 and N2. Immunochemical analysis of the mutant membranes with subunit-specific antibodies showed that the peripheral subunits were not assembled. Trans-complementation of the mutant strains by a native nuoI gene restored the wild-type phenotypes. In the C74S mutant, a limited amount of NuoI subunit still bound to the membraneous domain of Complex I, which is an indication that NuoI directly interacts with this domain. All these results clearly show that NuoI plays a critical role in the connection between the membraneous domain and the peripheral domain of Complex I.
编码荚膜红细菌复合体I中类铁氧化还原蛋白亚基（等同于牛TYKY亚基的一个亚基）的nuoI基因通过同源重组发生了突变。一个缺失nuoI的突变体（ΔnuoI突变体）和一个将半胱氨酸74替换为丝氨酸的点突变体（C74S突变体）均被证明在复合体I活性方面完全缺失。这些菌株在厌氧光合条件下无法生长。它们的细胞质膜还表现为不存在归属于铁硫簇N1和N2的特定电子顺磁共振信号。用亚基特异性抗体对突变体质膜进行免疫化学分析表明，外周亚基未组装。用天然nuoI基因对突变菌株进行反式互补恢复了野生型表型。在C74S突变体中，仍有少量NuoI亚基与复合体I的膜结构域结合，这表明NuoI直接与该结构域相互作用。所有这些结果清楚地表明，NuoI在复合体I的膜结构域和外周结构域之间的连接中起关键作用。
