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)

An 8.9-kb segment with hydrogenase genes from the cyanobacterium Anabaena variabilis has been cloned and sequenced. The sequences show homology to the methyl-viologen-reducing hydrogenases from archaebacteria and, even more striking, to the NAD(+)-reducing enzymes from Alcaligenes eutrophus and Nocardia opaca as well as to the NADP(+)-dependent protein from Desulfovibrio fructosovorans. The cluster from A. variabilis contains genes coding for both the hydrogenase heterodimer (hoxH and hoxY) and for the diaphorase moiety (hoxU and hoxF) described for the A. eutrophus enzyme. In A. variabilis the gene cluster is split by two open reading frames (between hoxY and hoxH and between hoxU and hoxY, respectively), and a probably non-coding 0.9-kb segment in an unusual way. The hoxH partial sequence from Anabaena 7119 and Anacystis nidulans was amplified by PCR. Using the labeled segment from A. 7119 as probe, Southern analysis revealed homologous gene segments in the cyanobacteria A. 7119, Anabaena cylindrica, Anacystis nidulans and A. variabilis. The bidirectional hydrogenase from A. nidulans was purified and digests were sequenced. The amino acid sequences obtained showed partial identities to the amino acid sequences deduced from the DNA data of the 8.9-kb segment from A. variabilis. Therefore the 8.9-kb segment contains the genes coding for the bidirectional, reversible hydrogenase from cyanobacteria. Crude extracts from A. nidulans perform NAD(P)H-dependent H2 evolution corroborating the molecular biological demonstration of the NAD(P)(+)-dependent hydrogenase in cyanobacteria.
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PMID:Molecular biological analysis of a bidirectional hydrogenase from cyanobacteria. 758 54

NAD(P)H:quinone oxidoreductase1 (DT-diaphorase or NQO1) is a flavoprotein that promotes obligatory two-electron reduction of quinones, preventing their participation in redox cycling, oxidative stress, and neoplasia. NQO1 is ubiquitously expressed. However, a large amount of variation in NQO1 gene expression was noticed among various human tissues. NQO1 gene is upregulated in livers of hepatocarcinoma patients, and its expression is induced in response to a variety of compounds, including planar aromatic hydrocarbons, phenolic antioxidants/chemoprotectors, tumor promoters, and hydrogen peroxide. Deletion mutagenesis in the NQO1 gene promoter identified several cis-elements including antioxidant response element (ARE), xenobiotic response element, and AP2 element, which regulate the expression and induction of the NQO1 gene. Among these DNA elements, ARE is the most important cis-element required for high basal expression of the NQO1 gene in tumor tissues, as compared to the normal tissues of the same origin, and for its induction in response to xenobiotics and antioxidants. Nucleotide sequence analysis of the ARE indicated presence of three AP1/AP1-like elements and a GCA box. Mutational analysis indicated a requirement of two AP1/AP1-like elements arranged as inverse repeats at the interval of three base pairs for the ARE activity. The GCA box in the ARE was required for optimum basal and induced expression. ARE is a novel cis-element because a single AP1/AP1-like element did not stimulate gene expression in response to xenobiotics and antioxidants. Band shift and supershift assays identified Jun, Fos, and novel proteins in the hARE-nuclear protein complexes that mediate regulation of the NQO1 gene expression.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:NAD(P)H:quinone oxidoreductase1 (DT-diaphorase): expression, regulation, and role in cancer. 762 Feb 21

In the present study, we examined whether active oxygen formed in the process of 4-nitroquinoline 1-oxide (4NQO) reduction by DT-diaphorase could induce oxidative stress on the pulmonary nuclei. The rapid production of OH- radical-like species after the start of the 4NQO reduction was observed, and subsequent induction of nuclear lipid peroxidation occurred. In conjugation with this event, DNA damage estimated as DNA single strand breaks (DNA-SSB) increased in a time-dependent manner. The induction of this DNA damage was partially inhibited by mannitol or vitamin E treatment. These findings suggest that the active oxygen generated in the process of the 4NQO reduction can induce oxidative damage on the pulmonary nuclei.
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PMID:Active oxygen generated in the process of carcinogen metabolism can induce oxidative damage in nuclei. 762 Aug 30

This study describes characteristics of a human bladder cancer cell line, SCaBER/R, selected for resistance to a mitomycin C (MMC) analogue BMY 25067. The SCaBER/R cell line was isolated by repeated 24 h exposures of the parental cells to 0.09 microM BMY 25067 (IC90, 24 h drug exposure) over a period of about 180 days. Approximately 2.2-fold higher concentration of BMY 25067 was required to kill 50% of the SCaBER/R cell line compared with parental cells (p < 0.001). The IC20 and IC90 values for BMY 25067 were also significantly higher in the SCaBER/R cell line than in SCaBER. Unlike most MMC resistant cell lines, the SCaBER/R cell line displayed a marked cross-resistance to 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) and lacked cross-resistance to cisplatin, doxorubicin or VP-16. The SCaBER/R cell line also displayed a marked cross-resistance to the parent drug (MMC) and BMY 25282, another analogue of MMC. NADPH cytochrome P450 reductase activity, an enzyme implicated in bio-reductive activation of MMC, did not differ significantly in these cells. DT-diaphorase activity, another MMC activation enzyme, was significantly lower in the SCaBER/R cell line when compared to the SCaBER cells. These results suggest that relatively lower sensitivity of SCaBER/R cell line to MMC and BMY 25067 may result from impaired drug activation. Cellular levels of glutathione (GSH) and GSH-transferase (GST), which have been suggested to affect the cytotoxicity of MMC, were comparable in SCaBER and SCaBER/R cell lines. BMY 25067 induced DNA interstrand cross-links (DNA-ISC) could not be detected in either of the cell lines even at drug concentrations which produced a significant cell kill. These findings suggest that (a) cellular resistance to BMY 25067 in the SCaBER/R cell line may be due to impaired drug activation, and (b) the nature of the cytotoxic produced by BMY 25067 may be different from that of MMC.
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PMID:Characterization of a human bladder cancer cell line selected for resistance to BMY 25067, a novel analogue of mitomycin C. 765 43

A nitroreductase isolated and purified from Escherichia coli B has been demonstrated to have potential applications in ADEPT (antibody-directed enzyme prodrug therapy) by its ability in vitro to reduce dinitrobenzamides (e.g. 5-aziridinyl 2,4-dinitrobenzamide, CB 1954 and its bischloroethylamino analogue, SN 23862) to form cytotoxic derivatives. In contrast to CB 1954, in which either nitro group is reducible to the corresponding hydroxylamine, SN 23862 is reduced by the nitroreductase to form only the 2-hydroxylamine. This hydroxylamine can react with S-acetylthiocholine to form a species capable of producing interstrand crosslinks in naked DNA. In terms of ADEPT, SN 23862 has a potential advantage over CB 1954 in that it is not reduced by mammalian DT diaphorases. Therefore, a series of compounds related to SN 23862 has been synthesized, and evaluated as potential prodrugs both by determination of kinetic parameters and by ratio of IC50 against UV4 cells when incubated in the presence of prodrug, with and without the E. coli enzyme and cofactor (NADH). Results from the two studies were generally in good agreement in that compounds showing no increase in cytotoxicity in presence of enzyme and cofactor were not substrates for the enzyme. None of the analogues were activated by DT diaphorase isolated from Walker 256 carcinoma cells. For those compounds which were substrates for the E. coli nitroreductase, there was a positive correlation between kcat and IC50 ratio. Two compounds showed advantageous properties: SN 25261 (with a dihydroxypropylcarboxamide ring substituent) which has a more than 10-fold greater aqueous solubility than SN 23862 whilst retaining similar kinetic characteristics and cytotoxic potency; and SN 25084, where a change in the position of the carboxamide group relative to the mustard resulted in an increased cytotoxicity ratio and kcat compared with SN 23862 (IC50 ratios 214 and 135; kcat values of 75 and 26.4 sec-1, respectively). An analogue (SN 25507) incorporating both these structural changes had an enhanced kcat of 576 sec-1. This study elucidates some of the structural requirements of the enzyme and aids identification of further directions in the search for suitable prodrugs for an ADEPT nitroreductase system.
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PMID:Bioactivation of dinitrobenzamide mustards by an E. coli B nitroreductase. 766 63

A point mutation in the mRNA of NADP(H): quinone oxidoreductase 1 (NQO1, DT-diaphorase) is believed to be responsible for reduced enzyme activity in the adenocarcinoma BE cell line. The present study examined nine cultured human non-cancerous fibroblast cell strains, five of which were from members of a single cancer-prone family, which demonstrated widely varying activity levels of DT-diaphorase (41 - 3462 nmol min-1 mg-1 protein), to determine if genetic alteration of the NQO1 or NOQ2 gene was involved in determining enzyme activity. All cell strains expressed NQO1 and NQO2 mRNA as measured by a quantitative polymerase chain reaction amplification technique. No relationship was found between the level of mRNA expressed and the enzyme activity in the cells. Sequencing of the entire complementary DNA from the cell strains revealed only a single base substitution at nucleotide 609 in one allele encoding NQO1 in every cell strain from members of the cancer-prone family, except for one cell strain which expressed only the T at nucleotide 609 in both alleles. Subsequent examination of genomic DNA from 44 individuals revealed that this base substitution is present in approximately 50% of the population. The presence of the T at nucleotide 609 in the NQO1 locus does not appear to be directly causal for altered DT-diaphorase activity.
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PMID:Presence of a heterozygous substitution and its relationship to DT-diaphorase activity. 766 61

Purified DT-diaphorase [NAD(P)H (quinone acceptor) oxidoreductase (EC.1.6.99.2)] from Walker cells was used to investigate the reductive metabolism of 3-amino-1,2,4-benzotriazine-1,4-dioxide (SR 4233) under aerobic and anaerobic conditions. In the presence of NADPH, under aerobic conditions, HPLC analysis showed the four-electron reduction product 3-amino-1,2,4-benzotriazine (SR 4330) was the major reaction product. In contrast, anaerobically, the 2-electron reduction product 3-amino-1,2,4-benzotriazine-1-oxide (SR 4317) was the predominant metabolite. Anaerobic reduction of SR 4233 to the known metabolites SR 4317 and SR 4330, catalyzed by DT-diaphorase, was 3-fold higher than reduction under aerobic conditions. Anaerobically, approximately half of the substrate utilized could not be accounted for by the formation of known products. Aerobically, the majority of the SR 4233 lost could be accounted for by its conversion to SR 4317 and SR 4330. In Walker cells incubated with SR 4233 anaerobically, SR 4317 was the major metabolite formed. Dicoumarol (100 microM) had little effect on the rate of formation of this metabolite in this cell line or in a rat liver epithelial derived (JBJ) cell line. Dicoumarol did however partially reduce the induction of unscheduled DNA synthesis caused by SR 4233 in Walker cells but not in JB1 cells, suggesting the action of dicoumarol may be specific to Walker cells. It is concluded that DT-diaphorase plays only a minor role in the overall reduction of SR 4233 in the two cell lines studied.
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PMID:Metabolism of 3-amino-1,2,4-benzotriazine-1,4-dioxide (SR 4233) by purified DT-diaphorase under aerobic and anaerobic conditions. 767 76

Using 4 human cancer cell lines, the relevance of NAD(P)H: quinone oxidoreductase (DT-diaphorase) activity to mitomycin C (MMC)-induced cytotoxicity was investigated. KB cells (oral epidermoid carcinoma) had more than 4 times higher DT-diaphorase activity than PH101 (pancreatic cancer), SH 101 (gastric cancer), or K562 (myelogenous leukemia) cells. The sensitivity to MMC was greatest in KB cells. Concentrations causing 50% inhibition of cell growth (IC50 value: microgram/ml) by 30 min treatment with MMC were 0.4 in KB, 1.1 in PH101, 1.6 in SH 101, and 1.9 in K 562. Treatment with 1.5 micrograms/ml of MMC induced DNA total cross links, and the indices were 0.18 in KB, 0.10 in SH101, 0.09 in SH101, and 0.06 in K 562. When DT-diaphorase activity was inhibited by non-toxic dicoumarol (50 microM), DNA damage and cytotoxic activity induced by MMC were decreased in all cells examined. Especially in KB cells, it was remarkable. Since it was shown that the level of cellular DT-diaphorase activities were correlated with the responses to MMC, we suggest that bioreduction by DT-diaphorase may activate MMC.
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PMID:[Mitomycin C and its bioreduction: relevance of NAD(P)H: quinone oxidoreductase activity to mitomycin C-induced DNA damage and cytotoxicity]. 768 84

We have cloned and sequenced the mouse NMO1 cDNA, which encodes the NAD(P)H:menadione oxidoreductase [also called NAD(P)H:(quinone acceptor) oxidoreductase; quinone reductase; azo dye reductase; DT diaphorase; EC 1.6.99.2]. The cDNA is 1528 bp in length excluding the poly(A+) tail, and has 5' and 3' nontranslated regions of 108 bp and 595 bp, respectively. The deduced protein contains 274 amino acids, including the first methionine (M(r) = 30,959). The mouse NMO1 protein is: 94% similar to the rat NMO1 and 86.5% to the human NMO1 proteins; 49.3% identical to the human NQO2 protein; and < 20% similar to several dozen other proteins in the quinone oxidoreductase superfamily. Southern hybridization analysis of mouse DNA reveals that the Nmo1 gene is likely to span less than a total of 20 kb. The Nmo1 gene is highly inducible by 2,3,7,8,-tetrachlorodibenzo-p-dioxin (dioxin; TCDD) in mouse liver and mouse cell cultures. TCDD inducibility of NMO1 is detectable at 12 and 18 days of gestation, but markedly elevated at 1-3 weeks post partum as compared with the 6- and 12-week-old mouse. NMO1 mRNA levels are strikingly elevated in the untreated mouse hepatoma Hepa-1c1c7 mutant line c37 lacking CYP1A1 (aryl hydrocarbon hydroxylase) activity, and in the untreated 14CoS/14CoS mouse cell line having an 'oxidative stress response' caused by homozygous deletion of about 3800 kb on chromosome 7. Previous work and the data in this report show that the murine Nmo1 gene is regulated by three distinct mechanisms: CYP1A1 metabolism-dependent repression, Ah receptor-mediated induction by TCDD, and activation by the chromosome 7-mediated oxidative stress response.
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PMID:Mouse dioxin-inducible NAD(P)H: menadione oxidoreductase: NMO1 cDNA sequence and genetic differences in mRNA levels. 770 40

We have demonstrated for the first time that mitoplasts (i.e. mitochondria without outer membrane) were able to convert stilbene estrogen (diethylstilbestrol, DES) to reactive metabolites, which covalently bind to mitochondrial (mt)DNA. Depending on the cofactor used, mitochondrial enzymes catalyzed the oxidation and/or reduction of DES. DES was oxidized to DES quinone by peroxide-supported mitochondrial enzyme. A Lineweaver-Burk plot of rate of formation of DES quinone at various substrate concentrations yielded a Km of 33 microM and Vmax of 39 nmol/mg protein/min. The oxidation of DES to DES quinone by mitochondria was drastically decreased by known inhibitors of cytochrome P450. DES quinone was reduced to DES by mitoplasts in the presence of NADH. The Km and Vmax for the DES quinone reduction in the absence of mitoplasts and NADH were 3.2 microM and 5.6 nmol respectively. The reduction of DES quinone to DES by mitoplasts was significantly inhibited by inhibitors of cytochrome b5 reductase and diaphorase. DES quinone was also reduced to DES by pure diaphorase, a mitochondrial reducing enzyme, in the presence of NADH. The Km and Vmax for the DES quinone reduction by diaphorase were 9.0 microM and 4.3 nmol respectively. Under reaction conditions similar to oxidation of DES to DES quinone by mitoplasts, it was observed that mitochondrial metabolic products of DES were able to covalently bind to mtDNA. These data provide direct evidence of mitochondrial enzyme-catalyzed oxidation and reduction reactions of DES. In the cell, activation of DES in the mitochondria (the organelle in which mtDNA synthesis, mtDNA repair and transcription systems are localized) is of utmost importance, because an analogous in vivo mitochondrial metabolism of DES through covalent modifications in mitochondrial genome may produce instability in the mitochondrial genome of the cells. These modifications may in turn play a role in the development of DES-induced hepatocarcinogenicity.
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PMID:Mitochondrial enzyme-catalyzed oxidation and reduction reactions of stilbene estrogen. 772 71


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