THE NUOI SUBUNIT OF THE RHODOBACTER-CAPSULATUS RESPIRATORY COMPLEX-I (EQUIVALENT TO THE BOVINE TYKY SUBUNIT) IS REQUIRED FOR PROPER ASSEMBLY OF THE MEMBRANOUS AND PERIPHERAL DOMAINS OF THE ENZYME

The nuoI gene that encodes a ferredoxin-like subunit of the Rhodobacte r capsulatus Complex I (a subunit equivalent to the bovine TYKY subuni t) 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 photosynt hetic 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-spec ific antibodies showed that the peripheral subunits were not assembled . Trans-complementation of the mutant strains by a native nuoI gene re stored the wild-type phenotypes. In the C74S mutant, a limited amount of NuoI subunit still bound to the membraneous domain of Complex I, wh ich is an indication that NuoI directly interacts with this domain. Al l these results clearly show that NuoI plays a critical role in the co nnection between the membraneous domain and the peripheral domain of C omplex I.