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)

We report the identification of an NADH-dependent haem-degrading system in ox heart mitochondria. The activity was localized to the mitochondrial inner membrane, specifically associated with complex I (NADH:ubiquinone oxidoreductase). The mitochondrial NADH-dependent haem-degradation activity was highly effective and displayed a rate nearly 60% higher than that of the microsomal activity. The following observations suggested the enzymic nature of the activity: (i) haem degradation by complex I did not proceed upon exposure to elevated temperature and extremes of pH; (ii) it displayed substrate specificity; (iii) it was inhibited by a substrate analogue; and (iv) it showed a cofactor requirement. Moreover, the activity was distinctly different from the ascorbate-mediated haem-degradation activity. Also, complex I differed from the microsomal NADPH:cytochrome c (P-450) reductase inasmuch as the formation of an effective interaction with the microsomal haem oxygenase could not be detected. Addition of purified haem oxygenase to complex I neither influenced the rate of haem degradation nor resulted in the formation of biliverdin IX alpha. In contrast, addition of haem oxygenase to NADPH:cytochrome c (P-450) reductase enhanced the rate of haem degradation by nearly 8-fold, and more than 60% of the degraded haem could be accounted for as biliverdin IX alpha. The haem-degrading activity of complex I appeared to involve the activity of H2O2, as the reaction was inhibited by nearly 90% by catalase, and propentdyopents were detected as reaction products. Intact haemoproteins such as cytochrome c and myoglobin were not effective substrates. However, the haem undecapeptide of cytochrome c was degraded at a rate equal to that observed for haem. Haematohaem was degraded at a rate 50% lower than that observed for haem. It is suggested that the NADH-dependent haem-degradation system may have a biological role in the regulation of the concentration of respiratory haemoproteins and the disposition of the aberrant forms of the mitochondrial haemoproteins.
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PMID:Characterization of an NADH-dependent haem-degrading system in ox heart mitochondria. 312 Jun 97

A method has been developed for the histochemical demonstration of a variety of dehydrogenases in freeze-dried or fixed resin-embedded tissue. Seven dehydrogenases were studied. Lactate dehydrogenase, NADH dehydrogenase and NADPH tetrazolium reductase were all demonstrable in sections of paraformaldehyde-fixed resin-embedded tissue. Freeze-dried specimens were embedded, without fixation, in glycol methacrylate resin or LR Gold resin at either 4 degrees C or -20 degrees C. All the dehydrogenases except succinate dehydrogenase retained their activity in freeze-dried, resin-embedded tissue. Enzyme activity was maximally preserved by embedding the freeze-dried tissue specimens in glycol methacrylate resin at -20 degrees C. The dehydrogenases were accurately localized without any diffusion when the tissue sections were incubated in aqueous media. Addition of a colloid stabilizer to the incubating medium was not required. Freeze-drying combined with low-temperature resin embedding permits accurate enzyme localization without diffusion, maintenance of enzyme activity and excellent tissue morphology.
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PMID:Dehydrogenase enzyme histochemistry on freeze-dried or fixed resin-embedded tissue. 324 Sep 50

The yeast Candida parapsilosis possesses two routes of electron transfer from exogenous NAD(P)H to oxygen. Electrons are transferred either to the classical cytochrome pathway at the level of ubiquinone through an NAD(P)H dehydrogenase, or to an alternative pathway at the level of cytochrome c through another NAD(P)H dehydrogenase which is insensitive to antimycin A. Analyses of mitoplasts obtained by digitonin/osmotic shock treatment of mitochondria purified on a sucrose gradient indicated that the NADH and NADPH dehydrogenases serving the alternative route were located on the mitochondrial inner membrane. The dehydrogenases could be differentiated by their pH optima and their sensitivity to amytal, butanedione and mersalyl. No transhydrogenase activity occurred between the dehydrogenases, although NADH oxidation was inhibited by NADP+ and butanedione. Studies of the effect of NADP+ on NADH oxidation showed that the NADH:ubiquinone oxidoreductase had Michaelis-Menten kinetics and was inhibited by NADP+, whereas the alternative NADH dehydrogenase had allosteric properties (NADH is a negative effector and is displaced from its regulatory site by NAD+ or NADP+).
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PMID:The alternative respiratory pathway of the yeast Candida parapsilosis: oxidation of exogenous NAD(P)H. 326 91

Hepatocyte cytotoxicity caused by substituted benzoquinones was associated with increased cytosolic Ca2+ concentration. p-Benzoquinone-induced hepatotoxicity was enhanced when the hepatocytes were loaded with Ca2+ by preincubation with ATP. A similar order of potency of the substituted benzoquinones in releasing Ca2+ from isolated mitochondria and inducing hepatocyte cytotoxicity was found; in decreasing order, this was 2-Br-, unsubstituted-, 2-CH3-, 2,6-(CH3O)2-, 2,6-(CH3)2-, 2,5-(CH3)2-, 2,3,5-(CH3)3-, and 2,3,5,6-(CH3)4-benzoquinones (duroquinone). The cellular products of quinone metabolism, hydroquinones and glutathione conjugates, did not cause mitochondrial Ca2+ release. Benzoquinone-induced mitochondrial Ca2+ release was preceded by GSH conjugate formation and NAD(P)H oxidation but followed by mitochondrial swelling. With duroquinone, a slow GSH and NADPH oxidation preceded Ca2+ release, but GSH oxidation did not occur with Se-deficient mitochondria lacking glutathione peroxidase activity. Cyanide-insensitive respiration was also observed with duroquinone but not with benzoquinone, suggesting that duroquinone undergoes redox cycling. GSH was depleted by both arylation and oxidation with 2,6-(CH3O)2-, 2,6-(CH3)2-, 2,5(CH3)2-, and 2,3,5-(CH3)3-benzoquinones. Benzoquinone concentrations that totally depleted GSH did not cause Ca2+ release until intramitochondrial NAD(P)H was oxidized. Ca2+ release was also prevented when NAD(P)H generation was stimulated by the presence of isocitrate or 3-hydroxybutyrate. This suggests that mitochondrial Ca2+ release is associated with NAD(P)H oxidation catalyzed by NADH dehydrogenase with benzoquinone or by the glutathione peroxidase-glutathione reductase system with duroquinone.
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PMID:Quinone toxicity in hepatocytes: studies on mitochondrial Ca2+ release induced by benzoquinone derivatives. 342 29

Mitochondrial dysfunction in ischemic liver has been demonstrated to be due to decrease in the intramitochondrial level of ATP and the subsequent disruption of the proton barrier of the inner membrane (Watanabe, F., Hashimoto, T. and Tagawa, K. (1985) J. Biochem. 97, 1229-1234). In this study, another injury process, impairment of the electron-transfer system, which occurred during reoxygenation of ischemic liver, was studied during reperfusion of cold preserved liver and during cold incubation of isolated rat-liver mitochondria. The sites of the respiratory chain that were sensitive to peroxidative damage were ubiquinone-cytochrome c oxidoreductase and NADH-ubiquinone oxidoreductase. These enzymic activities decreased with increase in lipid peroxidation. Incubation of submitochondrial particles with t-butyl hydroperoxide or with an NADPH-dependent peroxidation system decreased the enzymic activities of the electron-transport system. These data strongly suggested that lipid peroxidation during reoxygenation of ischemic liver impaired the electron-transfer system. Thus, mitochondria of ischemic liver suffer from two different types of injury: increase in proton permeability during anoxia, and decrease in enzymic activities of the electron-transport system during reoxygenation.
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PMID:Peroxidative injury of the mitochondrial respiratory chain during reperfusion of hypothermic rat liver. 380 61

Though previously described as very low or absent in yeast, we find significant pyridine nucleotide transhydrogenation (NADPH + acetyl pyridine-NAD+----NADP+ + acetyl pyridine-NADH) activity in yeast extracts when assayed at pH 8-9, and describe here the subcellular distribution and separation of the various molecular forms contributing to the total activity in two yeast species. Gentle subcellular fractionation reveals transhydrogenase activity only in the cytosolic fraction of both Saccharomyces cerevisiae and Candida utilis while intact mitochondria and microsomes are without activity. On sucrose gradient centrifugation, this soluble cytosolic activity proves to be primarily in a high-molecular-weight (greater than 10(6)) band which has salmon-colored fluorescence on uv illumination. Sonication of the particulate subcellular fractions solubilizes substantial transhydrogenase activity from mitochondria of C. utilis (but not from S. cerevisiae) which on sucrose gradients consists of both high (greater than 10(6))- and low-molecular-weight active fractions, each with yellow-green fluorescence. Ammonium sulfate fractionation and sucrose gradient centrifugation of protein solubilized from whole yeast of both species by vigorous homogenization with glass beads confirms the presence and fluorescence of these various molecular weight forms. The relationship of these activities to other enzymatic activities (especially the mitochondrial external NADH dehydrogenase) is discussed.
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PMID:Pyridine nucleotide transhydrogenations in yeast. 390 68

The NADH and NADPH ferricyanide reductase activities present in mitochondrial NADH-CoQ reductase preparations have been studied utilizing two photoaffinity pyridine nucleotide analogues: arylazido-beta-alanyl NAD+ (A3'-O-[3-[N-(4-azido-2-nitrophenyl)amino]propionyl]NAD+) and arylazido-beta-alanyl NADP+ (N3'-O-[3-[N-(4-azido-3-nitrophenyl)amino]propionyl]NADP+). For the NADH-K3Fe(CN)6 reductase activity, arylazido-beta-alanyl NAD+ was found to be, in the dark, a competitive inhibitor with respect to both NADH and K3Fe(CN)6 with Ki,app values of 9.7 and 15.5 microM, respectively. In comparison the NADP+ analogue exhibited weak noncompetitive inhibitor activity for this reaction against both substrates. Upon photoirradiation arylazido-beta-alanyl NAD+ inhibited NADH-K3Fe(CN)6 reductase up to 70% in the presence of a 25-fold molar excess of analogue over the enzyme concentration. This photodependent inhibition could be prevented by the presence, during irradiation, of the natural substrate NADH. In contrast complex kinetic results were obtained with studies of the effects of the pyridine nucleotide analogues of NADPH-K3Fe(CN)6 reductase activity in the dark. Photoirradiation of either analogue in the presence of the enzyme complex resulted in an activation of NADPH-dependent activity. The possibility that the NADPH-K3Fe(CN)6 reductase activity of complex I represents a summation of the combined ferricyanide reductase activity of the NADPH-NAD+ transhydrogenase and NADH oxidoreductase is suggested.
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PMID:Studies of the ferricyanide reductase activities of the mitochondrial reduced nicotinamide adenine dinucleotide-ubiquinone reductase (complex I) utilizing arylazido-beta-alanyl NAD+ and arylazido-beta-alanyl NADP+. 392 31

A membrane-associated NADH dehydrogenase from beef neutrophils was purified to homogeneity, using detergent (cholate plus Triton X-100) extraction and chromatography on DEAE-Sepharose CL-6B, agarose-hexane-NAD, and hydroxylapatite. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed an apparent subunit molecular weight of 17,500, but the enzyme was highly aggregated (Mr greater than 450,000) in nondenaturing gels containing 0.1% Triton X-100. The protein band in nondenaturing gels was also stained for activity using NADH and nitro blue tetrazolium. The enzyme showed greatest electron acceptor activity with ferricyanide (100%), followed by cytochrome c (3.5%), dichloroindophenol (2.7%), and cytochrome b5 (0.34%). No activity was seen with oxygen. The Km values for NADH and ferricyanide were 18 and 9.5 microM, respectively, and NAD+ was a weak competitive inhibitor (Ki = 118 microM). No activity was seen with NADPH. No effects were seen with mitochondrial respiratory inhibitors such as azide, cyanide, or rotenone, but p-chloromercuribenzoate was strongly inhibitory and N-ethylmaleimide was weakly inhibitory. No free flavin was detectable in enzyme preparations. Based upon kinetic, physical, and inhibition properties, this NADH dehydrogenase differs from those previously described in microsomes and erythrocyte plasma membrane.
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PMID:NADH dehydrogenase from bovine neutrophil membranes. Purification and properties. 394 Oct 77

Monodehydroascorbate reductase (EC 1.6.5.4) was purified from cucumber fruit to a homogeneous state as judged by polyacrylamide gel electrophoresis. The cucumber monodehydroascorbate reductase was a monomer with a molecular weight of 47,000. It contained 1 mol of FAD/mol of enzyme which was reduced by NAD(P)H and reoxidized by monodehydroascorbate. The enzyme had an exposed thiol group whose blockage with thiol reagents inhibited the electron transfer from NAD(P)H to the enzyme FAD. Both NADH and NADPH served as electron donors with Km values of 4.6 and 23 microM, respectively, and Vmax of 200 mol of NADH and 150 mol of NADPH oxidized mol of enzyme-1 s-1. The Km for monodehydroascorbate was 1.4 microM. The amino acid composition of the enzyme is presented. In addition to monodehydroascorbate, the enzyme catalyzed the reduction of ferricyanide and 2,6-dichloroindophenol but showed little reactivity with calf liver cytochrome b5 and horse heart cytochrome c. The kinetic data suggested a ping-pong mechanism for the monodehydroascorbate reductase-catalyzed reaction. Cucumber monodehydroascorbate reductase occurs in soluble form and can be distinguished from NADPH dehydrogenase, NADH dehydrogenase, DT diaphorase, microsome-bound NADH-cytochrome b5 reductase, and NADPH-cytochrome c reductase by its molecular weight, amino acid composition, and specificity of electron acceptors and donors.
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PMID:Monodehydroascorbate reductase from cucumber is a flavin adenine dinucleotide enzyme. 405 27

1. Paraquat and diquat produce only a slight increase in the oxygen uptake of rat liver mitochondria, and it is likely that they do not penetrate the mitochondrial membrane. 2. In mitochondrial fragments inhibited by antimycin A or by Amytal, both substances stimulate oxygen uptake with NADH or beta-hydroxybutyrate as substrate but not with succinate. The NADH dehydrogenase of the respiratory chain appears to be involved, at a site only partially inhibited by Amytal. 3. An NADPH oxidase activity is stimulated in rat liver microsomes by diquat, and to a smaller extent by paraquat; diquat also causes an NADH oxidase activity to develop. The effect is not inhibited by carbon monoxide or p-chloromercuribenzoate, and it is probable that a flavoprotein is involved by a mechanism not requiring thiol groups. 4. One molecule of oxygen can oxidize two molecules of NADPH in the stimulated microsomal system, the hydrogen peroxide produced being broken down by a catalase activity in the microsomes. 5. Diquat can stimulate NADH oxidase and NADPH oxidase activity in the postmicrosomal soluble fraction; the enzyme involved may be DT-diaphorase. 6. The mechanism of these reactions and their significance in relation to the toxicity of the dipyridilium compounds are discussed.
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PMID:The action of paraquat and diquat on the respiration of liver cell fractions. 438 31


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