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:1.6.5.2 (NQO1)
6,196 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

DT diaphorase [NAD(P)H:quinone oxidoreductase] activity was measured in subcellular fractions from homogenates of striatum, frontal cortex, hippocampus, cerebellum, hypothalamus and substantia nigra. This flavoprotein, which by definition oxidizes dihydronicotinamide adenine dinucleotide and dihydronicotinamide adenine dinucleotide phosphate at equal rates and is completely inhibited by 10(-5) M dicoumarol, was found to constitute 80-90% of the total dihydronicotinamide adenine dinucleotide- and dihydronicotinamide adenine dinucleotide phosphate-reductase activities in all brain regions studied. Antibodies raised against purified cytosolic DT diaphorase from the rat liver cross-reacted with the brain enzyme and inhibited soluble DT diaphorase from striatum and cerebellum to 80-90%. Immunohistochemical studies with the same antibodies demonstrated the occurrence of DT diaphorase immunoreactivity in a population of neurons in the substantia nigra and ventral tegmental area. In some neurons there was a colocalization of DT diaphorase and tyrosine hydroxylase-like immunoreactivity. The dense network of DT diaphorase-immunoreactive fibres in the striatum disappeared along with the dopaminergic innervation after 6-hydroxydopamine lesion. DT diaphorase immunoreactivity was also found in Bergmann glia, astrocytes and tanycytes. No correlation appeared to exist between the localization of neuronal DT diaphorase immunoreactivity and the dihydronicotinamide adenine dinucleotide phosphate-diaphorase-like activity, as defined by tetrazolium salt staining, used as a marker for certain peptidergic and cholinergic neurons. However, in, for example, glial cells in the cerebellum, DT diaphorase might contribute or be responsible for the histochemical dihydronicotinamide adenine dinucleotide phosphate-diaphorase activity.
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PMID:Distribution of DT diaphorase in the rat brain: biochemical and immunohistochemical studies. 290 55

NADH-cytochrome b5 reductase is the predominant NADH-diaphorase found in the human neutrophil (Blood 62:152, 1983). Although this reductase segregates with the light membranes of nitrogen-cavitated neutrophils separated on Percoll gradients (which include the plasma membrane markers alkaline phosphatase and NADPH-oxidase), it is approximately 95% excluded from plasma membrane-enriched phagocytic vacuoles. The reductase constitutes approximately 5% of the light membrane fraction FAD-flavoprotein (14.8 +/- 5.5 pmol/mg protein) and was found in equimolar concentration with a high potential b cytochrome also present in this light membrane fraction and tentatively identified as cytochrome b5. Isolation of the reductase from human neutrophils was accomplished by Triton X-114 solubilization of the light Percoll gradient membranes, followed by temperature-dependent phase separation and then affinity chromatography on AMP-Sepharose. The active preparation contained 1.3 mol FAD/mol protein, migrated on sodium dodecyl sulfate-polyacrylamide gels as a single band corresponding to an apparent mol wt of 45,000 daltons, exhibited a pl of 5.7 on chromatofocusing and was obtained in greater than 70% yield, with an overall purification of almost 900-fold. The purified enzyme was characterized by a high specificity for NADH as electron donor (Km = 6.4 mumol/L v Km greater than 1.6 mmol/L for NADPH) and exhibited a maximal turnover of ca. 30,000 min-1 at 22 degrees C with either ferricyanide or cytochrome b5 (Km = 10 nmol/L) as electron acceptor. Although the physical characterization and biochemical properties described here demonstrate that this neutrophil NADH b5 reductase is similar to the corresponding liver and erythrocyte enzymes, its unique function in the neutrophil has yet to be determined.
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PMID:Purification and characterization of the human neutrophil NADH-cytochrome b5 reductase. 299 39

Enzymes catalyzing the two-electron reduction of quinones to hydroquinones are thought to protect the cell against quinone-induced oxidative stress. Using menadione as a substrate, carbonyl reductase, a cytosolic, monomeric oxidoreductase of broad specificity for carbonyl compounds, was found to be the main NADPH-dependent quinone reductase in human liver, whereas DT-diaphorase, the principal two-electron transferring quinone reductase in rat liver, contributed a very minor part to the quinone reductase activity of human liver. Carbonyl reductase from liver was indistinguishable from carbonyl reductase previously isolated from brain (B. Wermuth, J. biol. Chem. 256, 1206 (1981] on the basis of molecular weight, isoelectric point, immunogenicity, substrate specificity and inhibitor sensitivity. The purified enzyme from liver catalyzed the reduction of a great variety of quinones. The best substrates were benzo- and naphthoquinones with short substituents, and the K-region orthoquinones of phenanthrene, benz(a)anthracene, pyrene and benzo(a)pyrene. A long hydrophobic side chain in the 3-position of the benzo- and naphthoquinones and the vicinity of a bay area or aliphatic substituent (pseudo bay area) to the oxo groups of the polycyclic compounds decreased or abolished the ability of the quinone to serve as a substrate. Non-k-region orthoquinones of polycyclic aromatic hydrocarbons were more slowly reduced than the corresponding K-region derivatives. The broad specificity of carbonyl reductase for quinones is in keeping with a role of the enzyme as a general quinone reductase in the catabolism of these compounds.
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PMID:Carbonyl reductase provides the enzymatic basis of quinone detoxication in man. 308 21

A quantitative histochemical assay for NADPH-ferrihemoprotein (P450) reductase had been developed. For optimal activity, it is necessary to use a relatively electropositive tetrazolium salt such as neotetrazolium chloride as the final acceptor. The apparent Km of the reaction is 0.83 mM. Its specificity has been proven in two ways: (i) activity is increased selectively in the pericentral zone of liver from rats treated with phenobarbitone, an inducer of the reductase, though not in liver of rats injected with 3-methylcholanthrene, which induces NAD(P)H dehydrogenase; (ii) it is competitively inhibited by NADP+ (Ki = 1.50 mM) though unaffected by dicumarol, an inhibitor of NAD(P)H dehydrogenase activity. An NADP+ concentration ten times greater than the substrate concentration inhibits the histochemical reaction and the reaction in a microsomal fraction assayed biochemically to the same degree (70% inhibition). The amount of inhibition is independent of temperature, of the zone of the acinus and of the treatment of the animal. Continuous microdensitometric monitoring of the reaction product as it is formed has shown that the specific reaction is linear with incubation up to 10 min, thus allowing end-point measurements to be used for cytophotometric analysis.
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PMID:A quantitative histochemical study of NADPH-ferrihemoprotein reductase activity. 309 10

Ferredoxin-NADP reductase from Euglena gracilis Klebs var. Bacillaris Cori purified to apparent homogeneity, yields a typical 36 kDa and an unusual 15 kDa polypeptide on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, exhibits a typical flavoprotein spectrum, contains FAD, and catalyzes NADPH-dependent iodonitrotetrazolium-violet diaphorase, NADPH-specific ferredoxin-dependent cytochrome-c-550 reductase and NADPH-NAD transhydrogenase activities. Rabbit antibody to the purified FNR blocks these activities specifically and also blocks the iodonitrotetrazolium-violet diaphorase activity of Euglena chloroplast completely. The low iodonitrotetrazolium-violet diaphorase activity in the plastidless mutant, W10BSmL, is mitochondrial and is not specifically blocked by the ferredoxin-NADP reductase antibody. Dark-grown non-dividing (resting) wild-type Euglena cells show a 4-fold increase in ferredoxin-NADP reductase activity during greening at 970 lx. Half of the low ferredoxin-NADP reductase activity in dark-grown cells is initially soluble, but by the end of chloroplast development nearly all of the enzyme is membrane-bound. The binding of ferredoxin-NADP reductase on exposure to light correlates with the extent of thylakoid membrane formation. Immunoblots of wild-type extracts during greening indicate that the 15 kDa polypeptide increases in the same manner as the extent of reductase binding to thylakoid membranes.
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PMID:Purification, properties, and cellular localization of Euglena ferredoxin-NADP reductase. 312 Jul 72

N-acetyl-p-benzoquinone imine (NAPQI), a reactive metabolite of acetaminophen, has previously been shown to be toxic to hepatocytes freshly isolated from rat liver [Mol. Pharmacol. 28:306-311 (1985)] NAPQI arylates and oxidizes cellular thiols, and either one or both reactions may be important in the pathogenesis of cytotoxicity. Two dimethylated analogues of NAPQI, N-acetyl-3,5-dimethyl-p-benzoquinone imine (3,5-diMeNAPQI) and N-acetyl-2,6-dimethyl-p-benzoquinone imine (2,6-diMeNAPQI), were prepared to determine whether one reaction might be more damaging to cells than the other. Of the three quinone imines, the least potent cytotoxin to rat hepatocytes was 3,5-diMeNAPQI. However, the cytotoxicity of 3,5-diMeNAPQI was markedly enhanced by pretreatment of cells with 1,3-bis-(2-chloroethyl)-N-nitrosourea, which inhibits glutathione reductase. Reactions of 3,5-diMeNAPQI with GSH, both chemically and in hepatocytes, indicated that this quinone imine primarily oxidized thiols. These findings were corroborated by results of covalent binding experiments, which showed that radiolabeled 3,5-diMeNAPQI bound only to a small extent to hepatocyte proteins. On the other hand, 2,6-diMeNAPQI, the most potent cytotoxin of the three quinone imines that was investigated bound extensively to hepatocyte proteins. In addition, 2,6-diMeNAPQI reacted with GSH, both chemically and in hepatocytes, to form significant amounts of GSSG. Reduction products of NAPQI and its dimethylated analogues were not important contributors to cytotoxicity or GSSG formation based on the following results: 1) the quinone imines did not increase oxygen consumption by hepatocytes nor did they lead to oxygen uptake in solution; 2) dicoumarol, an inhibitor of the reductase, DT-diaphorase, had no effect on cytotoxicity caused by the quinone imines. Evidence for the involvement of ipso-adducts of the quinone imines in their reactions with cellular thiols is provided by results of investigations on the effects of DTT on the metabolism, covalent protein binding, and cytotoxic effects of the quinone imines.
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PMID:Comparative cytotoxic effects of N-acetyl-p-benzoquinone imine and two dimethylated analogues. 317 35

The in vivo effects of oral anticoagulant therapy with 4-hydroxycoumarins on various vitamin K-dependent enzyme systems in man were compared. In hepatic microsomes obtained from donors who has been treated with 4-hydroxycoumarins for more than 6 months, the vitamin K 2,3 epoxide reductase activity and the DTT-dependent vitamin K quinone reductase activity were diminished to 35% and 20% of the corresponding normal values. In the non-hepatic tissues, only a small decrease in vitamin K 2,3 epoxide reductase activity could be demonstrated, while no differences were found in the vitamin K quinone reductase activities. In none of the tissues a significant increase of noncarboxylated precursor proteins was observed, whereas also vitamin K hydroquinone-dependent carboxylase activities seemed to be unaffected by the anticoagulant treatment.
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PMID:The in vivo effects of oral anticoagulants in man: comparison between liver and non-hepatic tissues. 326 Apr 13

Yeast glutathione reductase exists in a single molecular form which exhibits preferred NADPH and weak NADH linked multifunctional activities. Kinetic parameters for the NADPH and NADH linked reductase, transhydrogenase, electron transferase and diaphorase reactions have been determined. The functional preference for the NADPH linked reductase reaction is kinetically related to the high catalytic efficiency and low dissociation constants for substrates. NADP+ and NAD+ may interact with two different sites or different kinetic forms of the enzyme. The active site disulfide and histidine are required for the reductase activity but are not essential to the transhydrogenase, electron transferase and diaphorase activities. Amidation of carboxyl groups and Co(II) chelation of glutathione reductase facilitate the electron transferase reaction presumably by encouraging the formation of an anionic flavosemiquinone.
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PMID:Multifunctional activities of yeast glutathione reductase. 329 44

Vitamin K and 3- (and/or 2)-hydroxy-2,3-dihydro-2-methyl-3-phytyl-1,4-naphthoquinone (hydroxyvitamin K) have been identified as metabolites of vitamin K 2,3-epoxide incubated with hepatocytes isolated from normal and warfarin-resistant rats. Dithiothreitol added to the extracellular medium differentially enhanced the formation of both metabolites: hydroxyvitamin K formation, almost undetectable in the absence of dithiothreitol, was particularly affected. Addition of the vitamin K 2,3-epoxide reductase inhibitors warfarin (5 to 100 microM) and brodifacoum (1 to 5 microM) to normal rat hepatocyte cultures produced a slight increase in hydroxyvitamin K formation and a marked inhibition of vitamin K formation. Brodifacoum was a weak inhibitor of hydroxyvitamin K formation at higher concentrations. Hepatocytes from warfarin-resistant rats catalyzed hydroxyvitamin K formation 1.5 to 2 times faster and vitamin K formation 1.5 to 2 times slower than did normal rat hepatocytes. The addition of warfarin to these cultures had no effect on epoxide metabolism to hydroxyvitamin K and only partially diminished metabolism to vitamin K. In contrast, brodifacoum (1 microM) addition produced 50% inhibition of hydroxyvitamin K formation and almost complete inhibition of vitamin K formation. These data suggest that in resistant, but not in normal rat hepatocytes, the vitamin K 2,3-epoxide reductase makes a significant contribution to hydroxyvitamin K formation. A second sulfhydryl-dependent pathway, present in both strains, is also involved in the formation of this metabolite. They also suggest that in resistant rats, warfarin inhibition of the vitamin K 2,3-epoxide reductase, and presumably the sulfhydryl-dependent vitamin K reductase, is incomplete and independent of concentration.
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PMID:Normal and warfarin-resistant rat hepatocyte metabolism of vitamin K 2,3-epoxide: evidence for multiple pathways of hydroxyvitamin K formation. 339 32

Vitamin K-dependent carboxylase activity has been demonstrated in the crude microsomal fraction of the intima of bovine aortae. The procedure for the isolation of vessel wall carboxylase is a slight modification of the general preparation procedure for tissue microsomes. The highest activity of the non-hepatic enzyme was observed at 25 degrees C and hardly any NADH-dependent vitamin K reductase could be demonstrated. The optimal reaction conditions for both vessel wall as well as liver carboxylase were similar: 0.1 M-NaCl/0.05 M-Tris/HCl, pH 7.4, containing 8 mM-dithiothreitol, 0.4% 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulphonic acid (CHAPS), 0.4 mM-vitamin K hydroquinone and 2 M-(NH4)2SO4. Warfarin inhibits the hepatic and non-hepatic carboxylase/reductase enzyme complex more or less to a similar degree. We have measured the apparent Km values for the following substrates: Phe-Leu-Glu-Glu-Leu ('FLEEL'), decarboxylated osteocalcin, decarboxylated fragment 13-29 from descarboxyprothrombin and decarboxylated sperm 4-carboxyglutamic acid-containing (Gla-)protein. The results obtained demonstrated that liver and vessel wall carboxylase may be regarded as isoenzymes with different substrate specificities. The newly discovered enzyme is the first vitamin K-dependent carboxylase which shows an absolute substrate specificity: FLEEL and decarboxylated osteocalcin were good substrates for vessel wall carboxylase, but decarboxylated fragment 13-29 and decarboxylated sperm Gla-protein were not carboxylated at all.
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PMID:Isolation and partial characterization of a vitamin K-dependent carboxylase from bovine aortae. 349 40


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