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)

The system involved in the reduction of 2-[4'-di(2''-bromopropyl) aminophenylazolbenzoic acid (CB10-252), an agent designed for treating primary liver cell cancer, has been demonstrated to be localised mainly in the 108 000 X g supernatant fraction of rat liver homogenate. It is also present in other organs particularly in the spleen. DAB-azoreductase as shown previously is present almost entirely in the microsomal fraction and is found in high concentration only in liver. The pH maximum for CB10-252-azoreductase implying the importance of the 2'-carboxyl group in determining substrate specificity. The use of enzyme inhibitors and other additives showed that CB10-252 WAS NOT AXANTHINE OXIDASE OR DIHYDROFOLATE REDUCTASE. Its activity was not affected by carbon monoxide, phenobarbitone (PB), or 3-methylcholanthrene (MC) pretreatment. Enhancement of the activity by ferrous ions and FAD indicated that at least part of the reduction system could involve a flavoprotein with FAD as the prosthetic group. The activity of CB10-252-azoreductase and methylred-azoreductase was reduced by menadione (vitamin K3), cyanide and propylgallate. A diaphorase preparation from pig heart reduced both CB10-252 and methylred with both NADPH- and NADH-generating systems.
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PMID:Some characteristics of two azoreductase systems in rat liver. Relevance to the activity of 2-[4'-di(2"-bromopropyl)-aminophenylazo]benzoic acid (CB10-252), a compound possessing latent cytotoxic activity. 0 Jan 49

The distribution and relative activities of some dehydrogenases and diaphorases in the rat's carotid body and ganglion cervicale superior were studied histochemically and cytophotometrically. The treatment of rats by the medium with decreased oxygen and increased carbon dioxide contents, a reliable rise of NAD--diaphorase activities was found in the interlobe tissue and (less expressed and late appearing) in the principal cells. In contrast, such changes were not observed in the ganglion cervicale superioir, that have not a special chemoreceptory function. It is supposed that these changes in enzyme activities in the carotid body is due to the chemoreceptory function of the intralobe tissue containing the IInd type cells and "special" cells of sensitive nerve terminals.
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PMID:[Histochemical study of the chemoreceptor function of the carotid body in comparison with the superior cervical ganglion]. 69 2

Pyruvate:NADP+ oxidoreductase from Euglena gracilis, a homodimeric protein with a molecular weight of 309 kDa, is an iron-sulfur flavoenzyme that contains thiamin pyrophosphate (TPP). The functional structure of the enzyme was studied by a limited proteolysis experiment using trypsin. The evidence obtained shows that the enzyme consists of two functional domains, one of which contains an iron-sulfur cluster, which can be isolated as a homodimeric fragment of approximately 220 kDa by proteolysis. The other domain that contains FAD is released as a monomeric fragment of approximately 55 kDa. The pyruvate dehydrogenase reaction is still catalyzed by the large fragment when NADP+ is substituted by methyl viologen, while the small fragment retains a diaphorase-like electron-transfer activity from NADPH to MV. It is thus shown that pyruvate is oxidized in a CoA-dependent reaction to form CO2 and acetyl-CoA in the iron-sulfur domain, and that the two electrons formed are transferred to the FAD domain in which NADP+ is reduced. TPP is considered to be associated in the iron-sulfur domain. The NH2-terminal sequences of the enzyme and its proteolytic fragments reveal that the iron-sulfur domain occurs in the NH2-terminal side of the enzyme. For elucidation of the O2 instability of the enzyme, limited proteolysis was attempted in air. The tryptic fragment derived from the iron-sulfur domain, similar to the native enzyme, appears to be inactivated by direct contact with O2. In contrast, the FAD domain, when separated from the other domain, is quite stable in air, although the diaphorase activity decays when the native enzyme is exposed to O2.
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PMID:Pyruvate:NADP+ oxidoreductase from Euglena gracilis: limited proteolysis of the enzyme with trypsin. 191 Feb 87

Nitrofluoranthenes (NFs) are mutagenic and carcinogenic environmental pollutants found in incomplete combustion products and urban air particulate. We have studied both oxidative and reductive metabolism in vitro of different NF isomers mediated by subcellular rat liver fractions. Under aerobic conditions only ring hydroxylation of NFs by rat liver microsomes occurred and the isomeric position of the nitro group affected both the amount and the type of phenolic metabolites formed. Liver microsomes from 3-methylcholanthrene-induced rats were most effective in giving ring hydroxylated 7- and 8-nitrofluoranthene, whereas liver microsomes from phenobarbital-pretreated rats were the most active in metabolizing 1- and 3-nitrofluoranthene. Under anaerobic conditions, only reduction of NFs mediated by both cytosolic and microsomal rat liver enzymes occurred. Cofactor requirements and inhibition experiments indicated that the reductase activity in rat liver cytosolic fractions could be ascribed to DT-diaphorase, aldehyde oxidase and/or other unknown enzymes. The microsomal reductase activity was inhibited by oxygen, carbon monoxide, 2-diethylaminoethyl-2,2-diphenylvalerate hydrochloride and n-octylamine, and slightly by cytochrome c; flavin mononucleotide greatly enhanced this activity. 3-Nitrofluoranthene microsomal nitroreductase activity was increased by phenobarbital rat pretreatment and this increment correlated well with the content of cytochrome P450. These results indicate a participation of cytochrome P450 in the reductive metabolism of NFs by rat liver microsomes.
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PMID:Characterization of oxidative and reductive metabolism in vitro of nitrofluoranthenes by rat liver enzymes. 230 47

SR 4233 (3-amino-1,2,4-benzotriazine-1,4-dioxide) is a novel benzotriazine di-N-oxide which shows unusually high selective toxicity towards hypoxic cells, probably as a result of reductive bioactivation. Using an HPLC assay for the parent drug and its 2- and 4-electron reduction products (SR 4317 and SR 4330, respectively), we have examined the enzymology of SR 4233 reductive metabolism in vitro using a variety of different enzyme preparations. SR 4233 was converted extremely rapidly to SR 4317 under N2 by mouse liver microsomes, and showed a marked preference for NADPH over NADH as a reduced cofactor. The reaction was inhibited completely in air and boiled preparations. It was also inhibited by 78-86% in carbon monoxide (CO), implicating cytochrome P-450 as the major microsomal SR 4233 reductase. The kinetics of reductive metabolism of SR 4233 to SR 4317 by mouse liver microsomes conformed to Michaelis-Menten kinetics, with a Km of 1.4 mM and a Vmax of 950 nmol/min/mg protein. SR 4233 reduction was also catalysed by mouse liver cytosol under N2. However, rates were markedly slower than for microsomes and showed an equal dependency on NADH and NADPH. The cytosolic enzymes aldehyde oxidase and xanthine oxidase both catalysed SR 4233 reduction to SR 4317 under N2. Purified buttermilk xanthine oxidase also catalysed this reaction. In contrast to other enzyme preparations, DT-diaphorase from Walker 256 tumour cells reduced SR 4233 predominantly to SR 4330, and this reaction occurred under aerobic conditions. These data illustrate that SR 4233 is reduced rapidly by a wide variety of reductases. We propose that the therapeutic selectivity of SR 4233 will be controlled by the relative expression of reductases in tumour versus normal tissues, and in particular by the differential participation of putative activating versus detoxifying enzymes.
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PMID:Enzymology of the reductive bioactivation of SR 4233. A novel benzotriazine di-N-oxide hypoxic cell cytotoxin. 234 70

An enzymatic assay method for the determination of urinary formic acid is described. Formic acid in urine was cleaved to carbon dioxide and water by formic acid dehydrogenase, whereby NAD+ was converted to NADH, which reacted with INT (p-iodonitrotetrazolium violet) in the presence of NAD-diaphorase. The color thus produced was determined at 500 nm. In addition, a simple gas chromatographic method of urinary formic acid is described, in which head space gas of formic acid methylester was applied into the wide bore column. The urinary formic acid concentrations by the enzymatic method agreed well with that by the gas chromatographic method. A simple gas chromatographic method for urinary methanol assay is also described. Acetonitrile was added to an equal volume of urine containing methanol. After centrifugation, the supernatant was injected into gas chromatography (GC). The peaks of urinary methanol and ethanol were separated by GC. Formic acid and methanol in urine of unexposed healthy subjects and workers exposed to methanol were analyzed by the colorimetric and gas chromatographic methods. Geometric mean concentrations of urinary formic acid and methanol in the healthy subjects were 7.82 mg/g creatinine and 1.34 mg/l, respectively. The concentration ratio of formic acid to methanol in the urine of the workers exposed to methanol was calculated to be 3.67 +/- 2.10, which agreed with the ratio under a controlled exposure experiment. A slower excretion of formic acid than that of methanol in the urine of a volunteer was also observed.
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PMID:Enzymatic assay of formic acid and gas chromatography of methanol for urinary biological monitoring of exposure to methanol. 234 46

Hepatocytes isolated from phenobarbital (PB)-pretreated and naive male Sprague-Dawley rats were incubated with menadione under one of three oxygen conditions (0, 21, or 95% oxygen) for 3 hr. During this time, samples were drawn and assayed for lactate dehydrogenase release and trypan blue exclusion as indices of cytotoxicity. Neither parameter indicated any significant difference in menadione-induced cytotoxicity between naive and PB-pretreated hepatocytes. Likewise, no difference was observed between hepatocytes incubated in 21% versus 95% O2. Consistent with the oxyradical hypothesis of menadione-induced cytotoxicity, hepatocytes incubated under 0% O2 (95:5; N2:CO2) did not exhibit any menadione cytotoxicity. Hepatic microsomes prepared from PB-pretreated rats exhibited a threefold increase in NADPH cytochrome P450 reductase activity over those of controls. Menadione-stimulated superoxide (O2-) production was twofold higher in PB pretreated versus naive liver microsomes. However, PB pretreatment failed to produce an increase in O2- production in intact hepatocytes or in hepatocytes disrupted by sonication. The failure of PB pretreatment to increase menadione-induced cytotoxicity and superoxide production in either intact or sonicated hepatocytes suggests that a concomitant cytoprotective mechanism is induced as well. The data further indicate that the cytoprotective elements are located in a nonmicrosomal fraction of the cell. In support of this, we observed PB-induced increases in glutathione levels, glutathione reductase, and DT-diaphorase activities. These findings indicate that PB-induced enhancements of the hepatocellular cytoprotective mechanisms collectively compensate for the increased redox cycling mechanism, resulting in a mitigation of the anticipated increased hepatocellular cytotoxicity of menadione.
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PMID:Phenobarbital-induced cytosolic cytoprotective mechanisms that offset increases in NADPH cytochrome P450 reductase activity in menadione-mediated cytotoxicity. 254 42

Retinyl acetate, 13-cis-retinoic acid (13cisRA), and N-(4-hydroxyphenyl)-retinamide (4HPR) were assayed for their in vivo effects on hepatic levels of cytochrome P450, cytosolic glutathione-S-transferase, and quinone reductase. When given p.o. to Sprague-Dawley rats, all of the retinoids caused significant suppression in the levels of arylhydrocarbon hydroxylase, yet 13cisRA and 4HPR caused elevations in cytosolic levels of quinone reductase and glutathione-S-transferase, respectively. Scans of sodium dodecyl sulfate-polyacrylamide gels of microsomal proteins from the livers of retinoid-dosed animals showed changes in both the intensities and the number of stained bands. For microsomes from 13cisRA-dosed animals, there were additional changes in the absorption maximum of the carbon monoxide and octylamine difference spectra. There was, compared to controls, a 62% reduction in the NADPH-dependent binding of (+)-7S-trans-7,8-dihydro[7-14C]benzo(a)pyrene-7,8-diol to microsomal proteins from 13cisRA-dosed animals. Fluorography of the sodium dodecyl sulfate-polyacrylamide gels showed that the major reduction in metabolite binding occurred in the Mr 50,000 region of the gel. The reduction in the NADPH-dependent binding of (+)-7S-trans-7,8-dihydro[7-14C]benzo(a)pyrene-7,8-diol to microsomal proteins in vitro and the reduction in hepatic arylhydrocarbon hydroxylase levels correlated with a reduction in the in vivo binding of benzo(a)pyrene to rat liver DNA. Animals dosed for 7 days with 13cisRA, retinyl acetate, or 4HPR showed a 38, 27, and 40% reduction in binding of benzo(a)pyrene to liver DNA and a 29, 32, and 21% reduction in binding to stomach DNA, respectively, when the carcinogen was administered on the eighth day, and the tissues were harvested 24 h later. Binding to lung DNA was reduced by 23 and 11%, respectively, in the 13cisRA- and 4HPR-dosed rats. No differences were observed in binding to kidney. Thus, retinoids, by altering the metabolism of carcinogens, could influence the initiation stage of carcinogenesis.
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PMID:Effects of retinoids on metabolizing enzymes and on binding of benzo(a)pyrene to rat tissue DNA. 362 Nov 88

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

Reductive metabolism of carcinogenic 1-nitropyrene by rat liver microsomes and reconstituted cytochrome P-450 systems was investigated. Under the nitrogen atmosphere, 1-aminopyrene was the only detected metabolite of 1-nitropyrene. The reductase activity in liver 105,000 X g supernatant fraction was ascribed to DT-diaphorase, aldehyde oxidase, and other unknown enzyme(s) from the results of cofactor requirements and inhibition experiments. The microsomal reductase activity was inhibited by oxygen, carbon monoxide, 2,4-dichloro-6-phenylphenoxyethylamine, and n-octylamine. Flavin mononucleotide markedly enhanced the activity, and 2-diethylaminoethyl-2,2-diphenylvalerate hydrochloride also enhanced it, but slightly. The microsomal activity was induced by the pretreatment of rats with 3-methylcholanthrene, sodium phenobarbital, or polychlorinated biphenyl, and the increments of the activity correlated well with those of the specific contents of cytochrome P-450 in microsomes. The reductase activity could be reconstituted by NADPH-cytochrome P-450 reductase and forms of cytochrome P-450 purified from liver microsomes of polychlorinated biphenyl-induced rats. Among four forms of cytochrome P-450 examined, an isozyme P-448-IId which showed high activity in hydroxylation of benzo(a)pyrene catalyzed most efficiently the reduction of 1-nitropyrene. The results of this study indicate the central role of cytochrome P-450 in the reductive metabolism of 1-nitropyrene in liver microsomes.
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PMID:Participation of cytochrome P-450 in reductive metabolism of 1-nitropyrene by rat liver microsomes. 643 May 44


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