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 Co- and Ru-substituted derivatives of adrenal iron-sulfur protein (adrenodoxin) were prepared from its apoprotein in the presence of urea, dithiothreitol, Na2S, and metal ions. Both metal-substituted proteins had 2 g-atoms each of metal and labile sulfur per mole of protein. The Co derivative had optical absorption maxima at 257, 264, 470, and 1430 nm with shoulders at 275, 280, 300, and 380 nm. The molar extinction coefficient per Co atom was 2.200 M-1 cm-1 at 470 nm. The Ru derivative had a broad maximum at 500 nm with a molar extinction coefficient of approximately 100 M-1 cm-1 per Ru atom. The visible chromophore of the Co- and Ru-substituted proteins with mercurials revealed that the saturation levels are 8.6 and 8.4 mol of mercurial/mol of protein. The values agree with that of the native protein within experimental errors. The tyrosyl residue at position 82 displayed a broad anomalous emission at 335 and 331 nm for the Co- and Ru-substituted proteins, respectively, as well as in the case of the native protein. There was no electron paramagnetic resonance signal of the Co derivative in a wide magnetic field at 77 degrees K. Additionally, the Co and Ru derivatives had no enzymatic activity toward NADPH-cytochrome c reduction in the presence of adrenal diaphorase (adrenodoxin reductase). There was no indication that Mn, Ni, Cu, and Os are incorporated into the apoprotein in the presence of urea. Incorporation of Fe into the protein was examined in the presence of Co or Ru. In a system containing both Fe and Ru, Fe was exclusively incorporated into the protein. In contrast to this, the reaction products from a system containing both Fe and Co were found to consist of both Fe and Co derivatives at approximately equimolar quantity.
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PMID:Cobalt and ruthenium replacement for iron in adrenal iron-sulfur protein (adrenodoxin). Preparation and some properties. 23 19

Groups of male Wistar rats were fed semi-synthetic diets containing 0, 200 or 500 mg indole-3-carbinol (13C)/kg for 2, 7, 14 or 28 days. After 2 days, P-450 activities were already induced, but the isoenzyme pattern induced was different in the liver and the small intestine. Hepatic P4501A1, P4501A2 and P4502B1 apoprotein levels were dose-relatedly enhanced, whereas in the small intestine induced levels of P4502B1 and P4501A1 were detected but P4501A2 was not induced. Pentoxy- and ethoxyresorufin dealkylation (PROD and EROD) were dose-relatedly enhanced in the liver (5- and 7-fold, respectively, in the higher dose group) as well as in the small intestine (8- and 13-fold, respectively, at 500 mg 13C/kg diet). Testosterone 16 alpha- and 16 beta-hydroxylation in the small intestine were enhanced (6-9-fold) from day 2 onwards, but in the liver these activities were only slightly enhanced from day 7 onwards. Thus, the major forms induced in the liver appear to be P4501A1, P4501A2, P4502B1 and, to a lesser extent, P4503A, whereas in the small intestine all of the effects that were found are associated with only one cytochrome P-450, P4502B1. After 2 days I3C (500 mg/kg) induced glutathione S-transferase in the liver (1.3-fold) and small intestine (1.5-fold). Hepatic glucuronyl transferase (GT1) was induced (about 1.6-fold) after 7, 14 and 28 days. DT-diaphorase was induced in the liver (2.7-fold) and small intestine (1.5-fold) after 14 days of exposure to 500 mg I3C/kg diet. Treatment of rat hepatocytes with indole-3-acetonitrile and 3,3'-diindolylmethane, but not I3C and indole-3-carboxaldehyde, enhanced EROD activity and halved testosterone 16 alpha- and 2 alpha-hydroxylation. All four indoles slightly induced glutathione S-transferase in cultured hepatocytes. Thus, the in vitro studies suggest that the in vivo effects of I3C have to be attributed to indole-condensation products, such as 3,3'-diindolylmethane, but not to I3C itself.
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PMID:Effects of indole-3-carbinol on biotransformation enzymes in the rat: in vivo changes in liver and small intestinal mucosa in comparison with primary hepatocyte cultures. 152 33

Male Wistar rats were given semi-synthetic diets supplemented with 0, 2.5, 5 and 20% cooked Brussels sprouts for 2, 7, 14 or 28 days. The effects on several cytochrome P-450 enzymes and phase II enzymes (glutathione S-transferase (GST), glucuronyl transferases 1 and 2 (GT1 and GT2) and DT-diaphorase (DTD)) in the liver and small intestinal mucosa were investigated. From 2 days of exposure onwards Brussels sprouts induced P4501A2 and--to a lesser extent--P4501A1 apoprotein levels in the liver, whereas in the small intestine markedly enhanced P4502B apoprotein levels could be detected. No enhanced P4503A apoprotein levels were observed. The 5 and 20% sprouts diets increased the intestinal pentoxyresorufin depentylation (PROD, 4.5-9-fold), and the hydroxylation of testosterone at the 16 alpha- and 16 beta-site (2.6-4.2-fold) after 2 days of exposure. In addition, the 20% sprouts died also enhanced the intestinal ethoxyresorufin deethylation (EROD) activity (c. 5-fold), the hepatic EROD and PROD activities (c. 2-fold) and the formation of 6 beta-hydroxytestosterone (c. 1.6-fold); the formation of 2 alpha-hydroxytestosterone in the liver was decreased (to c. 70% of the control value). GST activity was induced both in the liver (5 and 20% diet) and intestine (20% diet only) throughout the experiment. The 20% sprouts diet enhanced the hepatic DTD and GT1 activities, whereas the GT2 activity was decreased. The induction of DTD in the small intestine after 2 days (2.5-3.2-fold with 5 and 20% sprouts diets, respectively) diminished during the experiment. These results indicate that dietary exposure to cooked Brussels sprouts for only 2 days can change the metabolic activities of several phase II enzymes and cytochrome P-450 enzymes, of which P4502B is the predominant form induced in the small intestine.
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PMID:Effects of cooked brussels sprouts on cytochrome P-450 profile and phase II enzymes in liver and small intestinal mucosa of the rat. 154 2

The potency of indole-3-carbinol (I3C) to form condensation products under acidic aqueous conditions was studied. After identifying a known dimer, 3,3'-diindolylmethane (DIM), we elucidated the structures of two trimers also found in acid reaction mixtures: 5,6,11,12,17,18-hexahydrocyclonona[1,2-b:4,5-b':7,8-b"]tri-indole (CTI), and 2,3-bis[3-indolylmethyl] indole (BII). The formation of these indole oligomers was shown to be pH dependent. The highest amounts of DIM and BII were formed in aqueous solutions having a pH value ranging from 4 to 5. No CTI could be detected at pH values above 4.5. In rats that received an oral dose of I3C we could detect DIM and BII in gastric contents, stomach tissue, small intestine and liver. No CTI could be detected in vivo after oral exposure to I3C. In in vitro experiments, using rat hepatocytes, the cytochrome P-450IA1 apoprotein level, 7-ethoxyresorufin O-deethylation activity (EROD) and DT-diaphorase activity (DTD) were markedly enhanced by DIM and CTI as well as BII.
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PMID:Structure elucidation of acid reaction products of indole-3-carbinol: detection in vivo and enzyme induction in vitro. 195 58

Biosynthesis of ferredoxin-NADP+ reductase in higher plants was investigated in relation with the mechanism of formation of the holoenzyme. The putative precursor of the flavoprotein, obtained after cell-free translation on a wheat germ extract primed with poly(A)-rich mRNA, was able to spontaneously bind free FAD, rendering a functional prereductase. The newly synthesized preholoenzyme showed diaphorase and cytochrome c reductase activities, an apparent molecular mass of 45 kDa, and contained FAD as the only flavin cofactor. It gave a positive reaction towards antisera against mature ferredoxin-NADP+ reductase. On the other hand, intracellular distribution of flavin-synthesizing enzymes indicates that FAD formation occurs in the cytoplasm; that is, in the same compartment as the site of reductase synthesis. On the basis of the preceding data a model is presented for the biosynthesis of the enzyme in vivo, involving conjugation of the apoprotein with FAD in the cytoplasm, followed by transport of the preholoreductase across the chloroplast envelope to reach its final destiny in the thylakoid membrane.
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PMID:Biosynthesis of ferredoxin-NADP+ oxidoreductase. Evidence for the formation of a functional preholoenzyme in the cytoplasmic compartment. 286 41

NAD(P)H: quinone-acceptor oxidoreductase (EC 1.6.99.2), also referred to as DT-diaphorase, is a flavoprotein that catalyzes the two-electron reduction of quinones and quinonoid compounds to hydroquinones, using either NADH or NADPH as the electron donor. Using an Escherichia coli expression system developed previously, we prepared three mutants of the rat liver quinone reductase. These mutants are Lys-113-His (K113H), Lys-113-Asp (K113D), and Lys-113-Ala (K113A). While the mutant K113H was readily purified using the same procedure as for the purification of the wild-type quinone reductase and found to have an activity similar to that of the wild-type enzyme, K113D and K113A were purified only in very small quantities, mainly in the form of apoprotein, and had very low activities. The results suggest that a positively charged amino acid at this position is important for the binding of the flavin adenine dinucleotide (FAD) prosthetic group. Flavin spectral studies of 6-mercapto-FAD-reconstituted mutants revealed that mutation at Lys-113 affects the protein environment around position-6 of the isoalloxazine ring.
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PMID:A site-directed mutagenesis study at Lys-113 of NAD(P)H:quinone-acceptor oxidoreductase: an involvement of Lys-113 in the binding of the flavin adenine dinucleotide prosthetic group. 763 39

The reaction mechanism of a 1,4-benzoquinone reductase from the wood-rotting basidiomycete Phanerochaete chrysosporium was investigated. The native, oxidized, FMN-containing enzyme was reduced quantitatively by NADH and the resulting reduced enzyme was reoxidized in the presence of one equivalent of 2,6-di-methoxy-1,4-benzoquinone (DMBQ). The stoichiometry of NADH oxidation versus DMBQ reduction is 1:1. The enzyme catalyzes the reduction of quinones to hydroquinones by a ping-pong steady-state mechanism. However, inhibition is observed at low NADH concentrations. Quinone products derived from the autooxidation of the unstable compounds 1,2,4-trihydroxybenzene and 5-chloro-2,3,4-trihydroxybenzene also appear to be substrates for the quinone reductase. The enzyme reduces the one-electron acceptors ferricyanide and ferricytochrome c (Cc3+) with rates of 58.4 and 0.08%, respectively, compared to DMBQ. The stoichiometry of NADH oxidation versus ferricyanide reduction is 1:2. In the presence of quinones the rates of Cc3+ and ferricyanide reduction are increased, owing to the nonenzymatic reduction of these acceptors by enzyme-generated hydroquinone products. Dicumarol and Cibacron blue are competitive inhibitors with respect to NADH, with Ki values of 2.1 and 0.30 microM, respectively. Reconstitution of the apoprotein with FMN yields a fully active enzyme at an FMN-to-protein ratio of 2:1, suggesting that the flavin content of the enzyme is two molecules of FMN per dimer.
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PMID:1,4-Benzoquinone reductase from basidiomycete Phanerochaete chrysosporium: spectral and kinetic analysis. 866 Jun 80

Ferredoxin and ferredoxin-NADP+ reductase are the two last partners of the photosynthetic electron-transfer chain, responsible for the final reduction of NADP+ to NADPH. Herein, we report the engineering and characterization of a novel protein molecule in which the electron-carrier protein (ferredoxin I) and the reductase (a flavoprotein) were covalently linked in a single polypeptide chain by gene fusion. The gene was obtained by joining the cDNAs encoding the respective proteins and subsequently by deleting the intervening sequence between them by site-directed mutagenesis. No extra amino acid residues were introduced between the C-terminus of ferredoxin I and the N-terminus of the flavoenzyme. The chimera was purified to homogeneity and characterized. The M(r) of the chimera apoprotein was 45,800 as determined by mass spectrometry, in agreement with the expected value of 45,846. Both flavin and iron-sulfur cluster were in 1:1 ratio with respect to the apoprotein. The chimera was found active as a diaphorase and, more interestingly, highly efficient as a cytochrome c reductase, without need for free ferredoxin addition in the assay medium. Several lines of evidence indicate that the ferredoxin and the reductase in the chimera assume a configuration quite similar to that in the dissociable physiological complex. Thus, the fusion protein could be a useful tool for studying the mechanism of protein-protein recognition and electron transfer in the ferredoxin-ferredoxin-NADP+ reductase system.
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PMID:A three-domain iron-sulfur flavoprotein obtained through gene fusion of ferredoxin and ferredoxin-NADP+ reductase from spinach leaves. 939 97

1. The herbicides butachlor (2-chloro-2',6',diethyl-N-[buthoxymethyl] acetanilide) and pretilachlor (2-chloro-2',6'-diethyl-N-[2-propoxyethyl] acetanilide) are widely used in Asia, South America, Europe and Africa. Isoprothiolane (diisopropyl-1,3-dithiolan-2-ylidenemalonate) is used as a fungicide and an insecticide in rice paddies. We administered these agrochemicals to the male rat and examined their effects on cytochrome P450 (P450), glutathione S-transferase (GST), UDP-glucuronosyltransferase (UDPGT), and NAD(P)H-quinone oxidoreductase 1 (NQO1)-related metabolism in the liver. 2. Administration of isoprothiolane, butachlor or pretilachlor to rat induced hepatic P4502B subfamily-dependent enzyme activities (pentoxyresorufin O-depentylation and testosterone 16 beta-hydroxylation) up to 271-413% of control, which coincided with the increase in expression levels of the P4502B apoprotein. 3. Activities of GST toward 1-chloro-2,4-nitrobenzene and 3,4-dichloronitrobenzene were slightly induced (127-133% of control) in the liver of the rat treated with these pesticides. On the other hand, marked elevations of UDPGT activities toward p-nitrophenol (164-281% of control) were observed. NQO1-related metabolism (menadione reductase activity) was also induced (123-176% of control) in the liver of rat treated with these agrochemicals. 4. These results indicate that some of the agrochemicals currently in use are capable of inducing phase I and II xenobiotic-metabolizing enzyme activities in an isozyme selective manner. The induction of these activities may disrupt normal physiologic functions related to these enzymes in exposed animals.
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PMID:Effects of the agrochemicals butachlor, pretilachlor and isoprothiolane on rat liver xenobiotic-metabolizing enzymes. 987 35

Cytochrome b(5) reductase (cb5r) catalyzes the transfer of reducing equivalents from NADH to cytochrome b(5). Utilizing an efficient heterologous expression system that produces a histidine-tagged form of the hydrophilic, diaphorase domain of the enzyme, site-directed mutagenesis has been used to generate cb5r mutants with substitutions at position 91 in the primary sequence. Arginine 91 is an important residue in binding the FAD prosthetic group and part of a conserved "RxY(T)(S)xx(S)(N)" sequence motif that is omnipresent in the "ferredoxin:NADP(+) reductase" family of flavoproteins. Arginine 91 was replaced with K, L, A, P, D, Q, and H residues, respectively, and all the mutant proteins purified to homogeneity. Individual mutants were expressed with variable efficiency and all exhibited molecular masses of approximately 32 kDa. With the exception of R91H, all the mutants retained visible absorption spectra typical of a flavoprotein, the former being produced as an apoprotein. Visible absorption spectra of R91A, L, and P were red shifted with maxima at 458 nm, while CD spectra indicated an altered FAD environment for all the mutants except R91K. Fluorescence spectra showed a reduced degree of intrinsic flavin fluorescence quenching for the R91K, A, and P, mutants, while thermal stability studies suggested all the mutants, except R91K, were somewhat less stable than the wild-type domain. Initial-rate kinetic measurements demonstrated that the mutants exhibited decreased NADH:ferricyanide reductase activity with the R91P mutant retaining the lowest activity, corresponding to a k(cat) of 283 s(-1) and a K(NADH)(m) of 105 microM, when compared to the wild-type domain (k(cat) = 800 s(-1) K(NADH)(m) = 6 microM). These results demonstrate that R91 is not essential for FAD binding in cb5r; however, mutation of R91 perturbs the flavin environment and alters both diaphorase substrate recognition and utilization.
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PMID:Arginine 91 is not essential for flavin incorporation in hepatic cytochrome b(5) reductase. 1133 12


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