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)

FNR is a transcriptional regulator that controls gene expression in response to oxygen limitation in Escherichia coli. The NADH dehydrogenase II gene (ndh) is repressed by FNR under anaerobic conditions. Repression is not simply due to occlusion of the promoter (-35 and -10) region by FNR because adjacent pairs of FNR monomers were found to bind at two sites centred at -50.5 and -94.5 in the ndh promoter region without preventing RNA polymerase binding. However, contact between RNA polymerase and the -132 to -62 region of the non-coding strand of ndh DNA, and RNA polymerase-mediated open complex formation, were prevented by bound FNR. The upstream FNR-binding site (-94.5) was needed for efficient FNR-dependent repression of ndh transcription in vitro, and also for repression of an ndh-lacZ fusion in vivo. Anaerobic ndh repression may thus involve the binding of two pairs of FNR monomers upstream of the -35 region, which prevents essential RNA polymerase-DNA contacts in the upstream region as well as inhibiting RNA polymerase function by direct FNR interaction. Expression of the ndh-lacZ fusion in an fnr deletion strain was enhanced by anaerobic growth in rich medium or minimal medium supplemented with amino acids. Furthermore, two proteins (M(r) 12,000 and 35,000) which interact with and may activate transcription from the ndh promoter under these conditions were detected by gel retardation analysis. These putative amino acid-responsive activators may thus offset FNR-mediated repression and maintain a low level of anaerobic ndh expression for regulating the NAD+/NADH ratio during growth in rich media.
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PMID:Regulation of transcription at the ndh promoter of Escherichia coli by FNR and novel factors. 806 61

We isolated and characterized mutants defective in nuo, encoding NADH dehydrogenase I, the multisubunit complex homologous to eucaryotic mitochondrial complex I. By Southern hybridization and/or sequence analysis, we characterized three distinct mutations: a polar insertion designated nuoG::Tn10-1, a nonpolar insertion designated nuoF::Km-1, and a large deletion designated delta(nuoFGHIJKL)-1. Cells carrying any of these three mutations exhibited identical phenotypes. Each mutant exhibited reduced NADH oxidase activity, grew poorly on minimal salts medium containing acetate as the sole carbon source, and failed to produce the inner, L-aspartate chemotactic band on tryptone swarm plates. During exponential growth in tryptone broth, nuo mutants grew as rapidly as wild-type cells and excreted similar amounts of acetate into the medium. As they began the transition to stationary phase, in contrast to wild-type cells, the mutant cells abruptly slowed their growth and continued to excrete acetate. The growth defect was entirely suppressed by L-serine or D-pyruvate, partially suppressed by alpha-ketoglutarate or acetate, and not suppressed by L-aspartate or L-glutamate. We extended these studies, analyzing the sequential consumption of amino acids by both wild-type and nuo mutant cells growing in tryptone broth. During the lag and exponential phases, both wild-type and mutant cells consumed, in order, L-serine and L-aspartate. As they began the transition to stationary phase, both cell types consumed L-tryptophan. Whereas wild-type cells then consumed L-glutamate, glycine, L-threonine, and L-alanine, mutant cells utilized these amino acids poorly. We propose that cells defective for NADH dehydrogenase I exhibit all these phenotypes, because large NADH/NAD+ ratios inhibit certain tricarboxylic acid cycle enzymes, e.g., citrate synthase and malate dehydrogenase.
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PMID:Mutations in NADH:ubiquinone oxidoreductase of Escherichia coli affect growth on mixed amino acids. 815 82

The existence of an organo-specific (heart) external NADH dehydrogenase located on the outer face of the inner mitochondrial membrane has been recently proposed. We have studied the respiration on external NADH in rat and beef heart mitochondrial fractions: (i) by using different mitochondrial isolation procedures on the rat, we observed that the higher the criteria of quality toward classical substrate respiration of mitochondrial fractions, the lower the external NADH-linked respiration; (ii) by using an especially loosely fitting glass-Teflon homogenizer, we obtained rat heart mitochondrial fractions practically free from external NADH linked respiration and with the highest respiratory control ratio on glutamate plus malate respiration. In rat and beef heart mitochondrial fractions containing an external NADH respiration: (i) ethoxyformic anhydride used previously to distinguish internal and external NADH oxidation was shown not to be specific; (ii) external NADH-linked respiration (although associated to the normally functioning respiratory chain as was shown by the effects of classic respiratory inhibitors) did not lead to ADP phosphorylation while glutamate plus malate did; (iii) respiratory activity on glutamate plus malate and external NADH was totally additive and the oxidation corresponded to two separate cytochrome oxidase pools, indicating a total functional separation between the two respiratory systems; (iv) NAD+ addition stimulated states 3 and 4 glutamate plus malate respiration to the same extent, indicating the presence of an appreciable number of internal dehydrogenases accessible to external cofactors. These results show that external NADH-linked dehydrogenase activity, which is usually detectable in mammal heart mitochondrial fractions, is of artefactual origin.
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PMID:The organo-specific external NADH dehydrogenase of mammal heart mitochondria has an artefactual origin. 839 14

The 3-subunit iron-sulfur flavoprotein (NADH-artificial electron acceptor oxidoreductase) derived from complex I (EC 1.6.5.3) is rapidly and irreversibly inactivated in the presence of NADH. The rate of inactivation increases with a decrease of the enzyme concentration. The activities with ferricyanide, menadione and cytochrome c were lost synchronously during preincubation of the enzyme in the presence of NADH or dithionite under either aerobic or anaerobic conditions. The titration of the inactivation rate with the NADH/NAD+ pair suggests that reduction of a component with Em' = -325 mV (n = 2) is a prerequisite for a loss of the enzyme activity. Among the compounds tested only FMN and NAD+ were able to protect the enzyme against the reductive inactivation. NADH-induced loss of the enzyme activity in diluted solutions is accompanied with the synchronous appearance of a fluorescence characteristic for free FMN. It is concluded that the reduction of flavin leads to a strong decrease of FMN affinity to its specific binding site, and possible implications of the redox-dependent affinity changes in operation of NADH-ubiquinone reductase are discussed.
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PMID:Reductive inactivation of the mitochondrial three subunit NADH dehydrogenase. 839 15

1. We describe the acyl-CoA and acyl-carnitine esters which arise from the incubation of well-coupled State 3 rat skeletal-muscle mitochondrial fractions with [U-14C]hexadecanoate and [U-14C]hexadecanoyl-carnitine. 2. Acyl-CoA ester intermediates of chain length 16, 14, 12, 10 and 8 carbons were detected. 3. Although incubations were in steady state in respect of oxygen consumption, 14CO2 production and generation of acid-soluble radioactivity, quantitative analysis of acyl-CoA esters showed that steady state was not achieved in respect of all intermediates. 4. 3-Hydroxyacyl- and 2-enoyl-CoA and -carnitine esters were found under normoxic conditions. 5. Direct measurement of NAD+ and NADH shows that under identical incubation conditions our observations cannot be explained by gross perturbation of the [NAD+]/[NADH] ratio. 6. We hypothesize that there is a small pool of rapidly recycling NAD+ channelled between complex I of the respiratory chain and the newly described mitochondrial-inner-membrane-associated beta-oxidation trifunctional enzyme [Uchida, Izai, Orii and Hashimoto (1992) J. Biol. Chem. 267, 1034-1041].
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PMID:Intramitochondrial control of the oxidation of hexadecanoate in skeletal muscle. A study of the acyl-CoA esters which accumulate during rat skeletal-muscle mitochondrial beta-oxidation of [U-14C]hexadecanoate and [U-14C]hexadecanoyl-carnitine. 842 53

The steady-state kinetics of the NADH dehydrogenase activities of the mitochondrial NADH-ubiquinone oxidoreductase in the presence of one-electron acceptors, ferricyanide and hexammineruthenium(III), were studied. Similar to ferricyanide, hexammineruthenium was found to be an efficient electron acceptor for the enzyme in inside-out submitochondrial particles and isolated Complex I, but not in intact mitochondria. Qualitatively the same results were obtained using submitochondrial particles or isolated Complex I. Both hexammineruthenium(III) and ferricyanide reduction was rotenone-insensitive and showed no stimulation by the uncouplers in tightly coupled submitochondrial particles. In contrast to the NADH-ferricyanide oxidoreductase reaction which exhibits a double substrate inhibition behaviour, no inhibition of the reaction by either NADH or the electron acceptor was revealed in the NADH-hexammineruthenium(III) reductase reaction. The double-reciprocal plots 1/v vs. 1/[NADH] at various hexammineruthenium(III) concentrations gave a series of straight lines intercepting in the third quadrant, thus supporting the mechanism of the overall reaction in which the reduced enzyme-NAD+ complex is oxidized by the electron acceptor before NAD+ dissociation. The apparent KsNADH values equal to 1 x 10(-5) and 4 x 10(-5) M for submitochondrial particles and Complex I, respectively (27 degrees C, pH 8.0), were determined from the secondary KmNADH vs. V (at different acceptor concentrations) plot. The Ki values for the competitive inhibition of NADH oxidation by NAD+ were 1 x 10(-3) M and 2 x 10(-3) M for the respective enzyme preparations. The results obtained suggest that hexammineruthenium(III) interacts with the NADH-ubiquinone oxidoreductase at a single reaction site different from that for fericyanide.
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PMID:Kinetics of the mitochondrial NADH-ubiquinone oxidoreductase interaction with hexammineruthenium(III). 844 12

Abnormalities of mitochondrial energy metabolism may play a role in normal aging and certain neurodegenerative disorders. In this regard, complex I of the electron transport chain has received substantial attention, especially in Parkinson's disease. The conventional method for studying complex I has been quantitation of enzyme activity in homogenized tissue samples. To enhance the anatomic precision with which complex I can be examined, we developed an autoradiographic assay for the rotenone site of this enzyme. [3H]dihydrorotenone ([3H]DHR) binding is saturable (KD = 15-55 nM) and specific, and Hill slopes of 1 suggest a single population of binding sites. Nicotinamide adenine dinucleotide (NADH) enhances binding 4- to 80-fold in different brain regions (EC50 = 20-40 microM) by increasing the density of recognition sites (Bmax). Nicotinamide adenine dinucleotide phosphate also increases binding, but NAD+ does not. In skeletal muscle, heart, and kidney, binding was less affected by NADH. [3H]DHR binding is inhibited by rotenone (IC50 = 8-20 nM), meperidine (IC50 = 34-57 microM), amobarbitol (IC50 = 375-425 microM), and MPP+ (IC50 = 4-5 mM), consistent with the potencies of these compounds in inhibiting complex I activity. Binding is heterogeneously distributed in brain with the density in gray matter structures varying more than 10-fold. Lesion studies suggest that a substantial portion of binding is associated with nerve terminals. [3H]DHR autoradiography is the first quantitative method to examine complex I with a high degree of anatomic precision. This technique may help to clarify the potential role of complex I dysfunction in normal aging and disease.
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PMID:[3H]dihydrorotenone binding to NADH: ubiquinone reductase (complex I) of the electron transport chain: an autoradiographic study. 865 75

We report on the loss of mitochondrial nicotinamide adenine dinucleotides in human cultured cells along with cell culture and acidification of the culture medium. This was established both by the direct measurement of the decrease in the mitochondrial NAD content and by the alteration of the oxidative properties of the mitochondria. In situ, this loss could be reversed in less than 2 h by changing the culture medium or by readjusting the pH of the medium at physiological pH values. By studying the oxidative properties of intact, but NAD-depleted, mitochondria in digitonin-permeabilized cells, we found that a rapid influx of NAD could replenish the mitochondrial NAD pool. This allowed the restoration of an active NAD+-dependent substrate oxidation. Depletion of mitochondrial NAD in cells grown under quiescent conditions was further confirmed by fluorimetric measurement of mitochondrial NAD, as was the influx of NAD+ into the mitochondrial matrix. These data constitute the first evidence of rapid fluxes of NAD through mitochondrial membranes in animal cells. They also point to the possible confusion between a loss of mitochondrial NAD and a defect of respiratory chain complex I in the context of screening procedures for respiratory chain disorder in human.
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PMID:Fluxes of nicotinamide adenine dinucleotides through mitochondrial membranes in human cultured cells. 866 5

The effect of methylglyoxal on the oxygen consumption of mitochondria of both normal and leukaemic leucocytes was tested by using different respiratory substrates and complex specific artificial electron donors and inhibitors. The results indicate that methylglyoxal strongly inhibits mitochondrial respiration in leukaemic leucocytes, whereas, at a much higher concentration, methylglyoxal fails to inhibit mitochondrial respiration in normal leucocytes. Methylglyoxal strongly inhibits ADP-stimulated alpha-oxoglutarate and malate plus NAD+-dependent respiration, whereas, at a higher concentration, methylglyoxal fails to inhibit succinate and alpha-glycerophosphate-dependent respiration. Methylglyoxal also fails to inhibit respiration which is initiated by duroquinone and cannot inhibit oxygen consumption when the N,N,N', N'-tetramethyl-p-phenylenediamine by-pass is used. NADH oxidation by sub-mitochondrial particles of leukaemic leucocytes is also inhibited by methylglyoxal. Lactaldehyde, a catabolite of methylglyoxal, can exert a protective effect on the inhibition of leukaemic leucocyte mitochondrial respiration by methylglyoxal. Methylglyoxal also inhibits l-lactic acid formation by intact leukaemic leucocytes and critically reduces the ATP level of these cells, whereas methylglyoxal has no effect on normal leucocytes. We conclude that methylglyoxal inhibits glycolysis and the electron flow through mitochondrial complex I of leukaemic leucocytes. This is strikingly similar to our previous studies on mitochondrial respiration, glycolysis and ATP levels in Ehrlich ascites carcinoma cells [Ray, Dutta, Halder and Ray (1994) Biochem. J. 303, 69-72; Halder, Ray and Ray (1993) Int. J. Cancer 54, 443-449], which strongly suggests that the inhibition of electron flow through complex I of the mitochondrial respiratory chain and inhibition of glycolysis by methylglyoxal may be common characteristics of all malignant cells.
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PMID:Selective inhibition of mitochondrial respiration and glycolysis in human leukaemic leucocytes by methylglyoxal. 916 22

Considerable quantitative variations in the competitive inhibition of NADH oxidase activity of bovine heart submitochondrial particles (SMP) by different samples of NAD- were observed. ADP-ribose (ADPR) was identified as the inhibitory contaminating substance responsible for variations in the inhibition observed. ADPR competitively inhibits NADH oxidation with Ki values (25 degrees C, pH 8.0) of 26 microM, 30 microM, and 180 microM for SMP, purified Complex I and three-subunit NADH dehydrogenase (FP), respectively. ADPR decreases NADH-induced flavin reduction and prolongs the cyclic bleaching of FP during aerobic oxidation of NADH. Ki for inhibition of the rotenone-sensitive NADH oxidase in SMP by ADPR does not depend on delta mu H+. The initial rate of the energy-dependent NAD+ reduction by succinate is insensitive to ADPR. The inhibitor increases the steady-state level of NAD+ reduction reached during aerobic succinate-supported reverse electron transfer catalyzed by tightly coupled SMP. The results obtained are consistent with the proposal on different nucleotide-binding sites operating in the direct and reverse reactions catalyzed by the mitochondrial NADH-ubiquinone reductase.
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PMID:A competitive inhibition of the mitochondrial NADH-ubiquinone oxidoreductase (complex I) by ADP-ribose. 923 Sep 20


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