Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.6.5.3 (complex I)
8,901 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

LHON (Leber hereditary optic neuropathy) is a maternally inherited disease that leads to sudden loss of central vision at a young age. There are three common primary LHON mutations, occurring at positions 3460, 11778 and 14484 in the human mtDNA (mitochondrial DNA), leading to amino acid substitutions in mitochondrial complex I subunits ND1, ND4 and ND6 respectively. We have now examined the effects of ND6 mutations on the function of complex I using the homologous NuoJ subunit of Escherichia coli NDH-1 (NADH:quinone oxidoreductase) as a model system. The assembly level of the NDH-1 mutants was assessed using electron transfer from deamino-NADH to the 'shortcut' electron acceptor HAR (hexammine ruthenium), whereas ubiquinone reductase activity was determined using DB (decylubiquinone) as a substrate. Mutant growth in minimal medium with malate as the main carbon source was used for initial screening of the efficiency of energy conservation by NDH-1. The results indicated that NuoJ-M64V, the equivalent of the common LHON mutation in ND6, had a mild effect on E. coli NDH-1 activity, while nearby mutations, particularly NuoJ-Y59F, NuoJ-V65G and NuoJ-M72V, severely impaired the DB reduction rate and cell growth on malate. NuoJ-Met64 and NuoJ-Met72 position mutants lowered the affinity of NDH-1 for DB and explicit C-type inhibitors, whereas NuoJ-Y59C displayed substrate inhibition by oxidized DB. The results are compatible with the notion that the ND6 subunit delineates the binding cavity of ubiquinone substrate, but does not directly take part in the catalytic reaction. How these changes in the enzyme's catalytic properties contribute to LHON pathogenesis is discussed.
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PMID:Leber hereditary optic neuropathy mutations in the ND6 subunit of mitochondrial complex I affect ubiquinone reduction kinetics in a bacterial model of the enzyme. 1789 48

The Na+-translocating NADH:quinone oxidoreductase (Na+-NQR) from Vibrio cholerae is a membrane-bound, respiratory Na+ pump. Its NqrF subunit contains one FAD and a [2Fe-2S] cluster and catalyzes the initial oxidation of NADH. A soluble variant of NqrF lacking its hydrophobic, N-terminal helix (NqrF') was produced in V. cholerae wild type and nqr deletion strain. Under identical conditions of growth and induction, the yield of NqrF' increased by 30% in the presence of the Na+-NQR. FAD-containing NqrF' species with or without the FeS cluster were observed, indicating that assembly of the FeS center, but not insertion of the flavin cofactor, was limited during overproduction in V. cholerae. A comparison of these distinct NqrF' species with regard to specific NADH dehydrogenase activity, pH dependence of activity and thermal inactivation showed that NqrF' lacking the [2Fe-2S] cluster was less stable, partially unfolded, and therefore prone to proteolytic degradation in V. cholerae. We conclude that the overall yield of NqrF' critically depends on the amount of fully assembled, FeS-containing NqrF' in the V. cholerae host cells. The Na+-NQR is proposed to increase the stability of NqrF' by stimulating the maturation of FeS centers.
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PMID:The Na+-translocating NADH:quinone oxidoreductase (Na+-NQR) from Vibrio cholerae enhances insertion of FeS in overproduced NqrF subunit. 1828 89

The mitochondrial oxidative phosphorylation involves five multimeric complexes imbedded in the inner membrane: complex I (Nicotinamide Adenine Dinucleotide (NADH) quinone oxidoreductase), II (succinate dehydrogenase), III (ubiquinol cytochrome c oxido reductase or bc1 complex), IV (cytochrome c oxidase), and V (ATP synthase). These respiratory complexes are conserved from the yeast Saccharomyces cerevisiae to human with the exception of complex I, which is replaced by three NADH dehydrogenases in S. cerevisiae. Here, we provide several protocols allowing an exhaustive characterization of each yeast complex: this chapter describes procedures from mitochondria preparation to measurement of the activity of each complex and analysis of their subunit composition and provides information on the interactions between different complexes.
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PMID:Preparation of respiratory chain complexes from Saccharomyces cerevisiae wild-type and mutant mitochondria : activity measurement and subunit composition analysis. 1837 11

NADH-quinone oxidoreductase (complex I) in bovine heart mitochondria has a molecular weight of approximately 1 million Da composed of 45 distinct subunits. It is the largest energy transducing complex so far known. Bacterial complex I is simpler and smaller, but the essential redox components and the basic mechanisms of electron and proton translocation are the same. Over the past three decades, Ohnishi et al. have pursued extensive EPR studies near liquid helium temperatures and characterized most of the iron-sulfur clusters in complex I. Recently, Yakovlev et al. [G. Yakovlev, T. Reda, J. Hirst, Reevaluating the relationship between EPR spectra and enzyme structure for the iron-sulfur clusters in NADH:quinone oxidoreductase, Proc. Natl. Acad. Sci. U. S. A. 104 (2007) 12720-12725] challenged Ohnishi's group by claiming that there were EPR "misassignments" among clusters N4, N5 and N6b (in order to prevent confusion, we used current consensus nomenclature, as the nickname). They claimed that we misassigned EPR signals arising from cluster N5 to cluster N4, and signals from cluster N6b to cluster N4. They also proposed that cluster N5 has (4Cys)-ligands. Based on the accumulated historical data and recent results of our site-specific mutagenesis experiments, we confirmed that cluster N5 has (1His+3Cys)-ligands as we had predicted. We revealed that E. coli cluster N5 signals could be clearly detected at the sample temperature around 3 K with microwave power higher than 5 mW. Thus Hirst's group could not detect N5 signals under any of their EPR conditions, reported in their PNAS paper. It seems that they misassigned the signals from cluster N4 to N5. As to the claim of "misassignment" between clusters N4 and N6b, that was not a possibility because our mutagenesis systems did not contain cluster N6b. Therefore, we believe that we have not made any "misassignment" in our work.
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PMID:Were there any "misassignments" among iron-sulfur clusters N4, N5 and N6b in NADH-quinone oxidoreductase (complex I)? 1848 92

NADH:quinone oxidoreductase (complex I) plays a central role in cellular energy metabolism, and its dysfunction is found in numerous human mitochondrial diseases. Although the understanding of its structure and function has been limited, the x-ray crystal structure of the hydrophilic part of Thermus thermophilus complex I recently became available. It revealed the localization of all redox centers, including 9 iron-sulfur clusters and their coordinating ligands, and confirmed the predictions mostly made by Ohnishi et al. (Ohnishi, T., and Nakamaru-Ogiso, E. (2008) Biochim. Biophys. Acta 1777, 703-710) based on various EPR studies. Recently, Yakovlev et al. (Yakovlev, G., Reda, T., and Hirst, J. (2007) Proc. Natl. Acad. Sci. U. S. A. 104, 12720-12725) claimed that the EPR signals from clusters N4, N5, and N6b were misassigned. Here we identified and characterized cluster N5 in the Escherichia coli complex I whose EPR signals had never been detected by any group. Using homologous recombination, we constructed mutant strains of H101A, H101C, H101A/C114A, and cluster N5 knock-out. Although mutant NuoEFG subcomplexes were dissociated from complex I, we successfully recovered these mutant NuoCDEFG subcomplexes by expressing the His-tagged NuoCD subunit, which had a high affinity to NuoG. The W221A mutant was used as a control subcomplex carrying wild-type clusters. By lowering temperatures to around 3 K, we finally succeeded in detecting cluster N5 signals in the control for the first time. However, no cluster N5 signals were found in any of the N5 mutants, whereas EPR signals from all other clusters were detected. These data confirmed that, contrary to the misassignment claim, cluster N5 has a unique coordination with His(Cys)(3) ligands in NuoG.
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PMID:Iron-sulfur cluster N5 is coordinated by an HXXXCXXCXXXXXC motif in the NuoG subunit of Escherichia coli NADH:quinone oxidoreductase (complex I). 1860 33

The mitochondrial generation of reactive oxygen species (ROS) plays a central role in many cell signaling pathways, but debate still surrounds its regulation by factors, such as substrate availability, [O2] and metabolic state. Previously, we showed that in isolated mitochondria respiring on succinate, ROS generation was a hyperbolic function of [O2]. In the current study, we used a wide variety of substrates and inhibitors to probe the O2 sensitivity of mitochondrial ROS generation under different metabolic conditions. From such data, the apparent Km for O2 of putative ROS-generating sites within mitochondria was estimated as follows: 0.2, 0.9, 2.0, and 5.0 microM O2 for the complex I flavin site, complex I electron backflow, complex III QO site, and electron transfer flavoprotein quinone oxidoreductase of beta-oxidation, respectively. Differential effects of respiratory inhibitors on ROS generation were also observed at varying [O2]. Based on these data, we hypothesize that at physiological [O2], complex I is a significant source of ROS, whereas the electron transfer flavoprotein quinone oxidoreductase may only contribute to ROS generation at very high [O2]. Furthermore, we suggest that previous discrepancies in the assignment of effects of inhibitors on ROS may be due to differences in experimental [O2]. Finally, the data set (see supplemental material) may be useful in the mathematical modeling of mitochondrial metabolism.
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PMID:Oxygen sensitivity of mitochondrial reactive oxygen species generation depends on metabolic conditions. 1936 81

Tuberculosis is the leading cause of death due to a single infectious agent in the world and the emergence of multidrug-resistant strains prompted us to develop new drugs with novel targets and mechanism. Here, we screened a natural antibiotics library with Mycobacterium smegmatis membrane-bound dehydrogenases and identified polymyxin B (cationic decapeptide) and nanaomycin A (naphtoquinone derivative) as inhibitors of alternative NADH dehydrogenase [50% inhibitory concentration (IC(50)) values of 1.6 and 31 microg/ml, respectively] and malate: quinone oxidoreductase (IC(50) values of 4.2 and 49 microg/ml, respectively). Kinetic analysis on inhibition by polymyxin B showed that the primary site of action was the quinone-binding site. Because of the similarity in K(m) value for ubiquinone-1 and inhibitor sensitivity, we examined amino acid sequences of actinobacterial enzymes and found possible binding sites for L-malate and quinones. Proposed mechanisms of polymyxin B and nanaomycin A for the bacteriocidal activity were the destruction of bacterial membranes and production of reactive oxygen species, respectively, while this study revealed their inhibitory activity on bacterial membrane-bound dehydrogenases. Screening of the library with bacterial respiratory enzymes resulted in unprecedented findings, so we are hoping that continuing efforts could identify lead compounds for new drugs targeting to mycobacterial respiratory enzymes.
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PMID:Polymyxin B identified as an inhibitor of alternative NADH dehydrogenase and malate: quinone oxidoreductase from the Gram-positive bacterium Mycobacterium smegmatis. 1956 54

Gramicidin S and polymyxins are small cationic cyclic peptides and act as potent antibiotics against Gram-negative and Gram-positive bacteria by perturbing integrity of the bacterial membranes. Screening of a natural antibiotics library with bacterial membrane vesicles identified gramicidin S as an inhibitor of cytochrome bd quinol oxidase and an alternative NADH dehydrogenase (NDH-2) and polymyxin B as an inhibitor of NDH-2 and malate: quinone oxidoreductase. Our studies showed that cationic cyclic peptide antibiotics have novel molecular targets in the membrane and interfere ligand binding on the hydrophobic surface of enzymes. Improvement of the toxicity and optimization of the structures and clinical uses are urgently needed for their effective application in combating drug-resistant bacteria.
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PMID:Gramicidin S and polymyxins: the revival of cationic cyclic peptide antibiotics. 1970 17

Complex I (NADH:quinone oxidoreductase) is crucial to respiration in many aerobic organisms. In mitochondria, it oxidizes NADH (to regenerate NAD+ for the tricarboxylic acid cycle and fatty-acid oxidation), reduces ubiquinone (the electrons are ultimately used to reduce oxygen to water) and transports protons across the mitochondrial inner membrane (to produce and sustain the protonmotive force that supports ATP synthesis and transport processes). Complex I is also a major contributor to reactive oxygen species production in the cell. Understanding the mechanisms of energy transduction and reactive oxygen species production by complex I is not only a significant intellectual challenge, but also a prerequisite for understanding the roles of complex I in disease, and for the development of effective therapies. One approach to defining a complicated reaction mechanism is to break it down into manageable parts that can be tackled individually, before being recombined and integrated to produce the complete picture. Thus energy transduction by complex I comprises NADH oxidation by a flavin mononucleotide, intramolecular electron transfer from the flavin to bound quinone along a chain of iron-sulfur clusters, quinone reduction and proton translocation. More simply, molecular oxygen is reduced by the flavin, to form the reactive oxygen species superoxide and hydrogen peroxide. The present review summarizes and evaluates experimental data that pertain to the reaction mechanisms of complex I, and describes and discusses contemporary mechanistic hypotheses, proposals and models.
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PMID:Towards the molecular mechanism of respiratory complex I. 2002 15

Iron-sulfur cluster N2 of complex I (proton pumping NADH:quinone oxidoreductase) is the immediate electron donor to ubiquinone. At a distance of only approximately 7A in the 49-kDa subunit, a highly conserved tyrosine is found at the bottom of the previously characterized quinone binding pocket. To get insight into the function of this residue, we have exchanged it for six different amino acids in complex I from Yarrowia lipolytica. Mitochondrial membranes from all six mutants contained fully assembled complex I that exhibited very low dNADH:ubiquinone oxidoreductase activities with n-decylubiquinone. With the most conservative exchange Y144F, no alteration in the electron paramagnetic resonance spectra of complex I was detectable. Remarkably, high dNADH:ubiquinone oxidoreductase activities were observed with ubiquinones Q1 and Q2 that were coupled to proton pumping. Apparent Km values for Q1 and Q2 were markedly increased and we found pronounced resistance to the complex I inhibitors decyl-quinazoline-amine (DQA) and rotenone. We conclude that Y144 directly binds the head group of ubiquinone, most likely via a hydrogen bond between the aromatic hydroxyl and the ubiquinone carbonyl. This places the substrate in an ideal distance to its electron donor iron-sulfur cluster N2 for efficient electron transfer during the catalytic cycle of complex I.
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PMID:The role of a conserved tyrosine in the 49-kDa subunit of complex I for ubiquinone binding and reduction. 2011 74


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