The mitochondrial proton-translocating NADH-quinone oxidoreductase (complex I) is one of the four enzyme complexes in the respiratory chain. Complex I is composed of at least 46 different subunits with a total molecular mass of approximately 1Mda. It is recognized that complex I has the most intricate structure of the membrane-bound enzyme complexes. At present, high resolution 3D structures are not available. Low resolution EM analyses indicate that complex I has an L-shaped structure consisting of a membrane domain and a peripheral domain extending to the matrix. Complex I contains one FMN and eight FeS clusters as cofactors. These cofactors are considered to be located in the peripheral domain. This enzyme pumps protons from the matrix side to the cytoplasmic side and builds an electrochemical gradient across the inner-mitochondrial membrane. Recently, it has been reported that defects of complex I are involved in many human mitochondrial diseases. Evidently, studies of complex I are important both in the basic sciences and in medical research. In contrast to mitochondria, the respiratory chain of bacteria harbors a proton-translocating NADH-quinone oxidoreductase which, although quite similar to complex I in terms of cofactors, is much simpler in structure. This bacterial equivalent of complex I is referred to as NDH-1 and is made up of only 14 unlike subunits, all of which have homologues in complex I. Because of its structural simplicity, NDH-1 is a useful model system for studying the structure and function of complex I. The overall goal of this grant application is to elucidate the structure and the mechanism of action of the NDH-1/complex I. The studies planned for this grant period are as follows. (1) Characterization of FeS clusters, determination of the amino acid residues involved in coordination of the FeS clusters, and clarification of physiological roles of the FeS clusters. (2) Identification of the conserved amino acid residues in the membrane domain subunits which are essential for structure and function of the NDH-1. (3) Determination of subunit-subunit interaction of the NDH-1. (4) Identification of the subunit(s) and the amino acid residue(s) labeled by photoaffinity analogues of inhibitors specific to the NDH-1/complex I.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
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Physical Biochemistry Study Section (PB)
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Preusch, Peter C
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Scripps Research Institute
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Sinha, Prem Kumar; Castro-Guerrero, Norma; Patki, Gaurav et al. (2015) Conserved amino acid residues of the NuoD segment important for structure and function of Escherichia coli NDH-1 (complex I). Biochemistry 54:753-64
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Torres-Bacete, Jesus; Sinha, Prem Kumar; Sato, Motoaki et al. (2012) Roles of subunit NuoK (ND4L) in the energy-transducing mechanism of Escherichia coli NDH-1 (NADH:quinone oxidoreductase). J Biol Chem 287:42763-72
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Sinha, Prem Kumar; Nakamaru-Ogiso, Eiko; Torres-Bacete, Jesus et al. (2012) Electron transfer in subunit NuoI (TYKY) of Escherichia coli NADH:quinone oxidoreductase (NDH-1). J Biol Chem 287:17363-73
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Torres-Bacete, Jesus; Sinha, Prem Kumar; Matsuno-Yagi, Akemi et al. (2011) Structural contribution of C-terminal segments of NuoL (ND5) and NuoM (ND4) subunits of complex I from Escherichia coli. J Biol Chem 286:34007-14
Murai, Masatoshi; Yamashita, Tetsuo; Senoh, Mai et al. (2010) Characterization of the ubiquinone binding site in the alternative NADH-quinone oxidoreductase of Saccharomyces cerevisiae by photoaffinity labeling. Biochemistry 49:2973-80

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