Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: KEGG:D02011 (FAD)
 5,530 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Evidence is presented which suggests that the NAD(P)H-cytochrome c reductase component of nitrate reductase is the main site of action of the inactivating enzyme. When tested on the nitrate reductase (NADH) from the maize root and scutella, the NADH-cytochrome c reductase was inactivated at a greater rate than was the FADH2-nitrate reductase component. With the Neurospora nitrate reductase (NADPH) only the NADPH-cytochrome c reductase was inactivated. p-Chloromercuribenzoate at 50 muM, which gave almost complete inhibition of the NADH-cytochrome c reductase fraction of the maize nitrate reductase, had no marked effect on the action of the inactivating enzyme. A reversible inactivation of the maize nitrate reductase has been shown to occur during incubation with NAD(P)H. In contrast to the action of the inactivating enzyme, it is the FADH2-nitrate reductase alone which is inactivated. No inactivation of the Neurospora nitrate reductase was produced by NAD(P)H alone and also in the presence of FAD. The lack of effect of the inactivating enzyme and NAD(P)H on the FADH2-nitrate reductase of Neurospora suggests some differences in its structure or conformation from that of the maize enzyme. A low level of cyanide (0.4 mu M) markedly enhanced the action of NAD(P)H on the maize enzyme; Cyanide at a higher level (6 mu M) did give inactivation of the Neurospora nitrate reductase in the presence of NADPH and FAD. The maize nitrate reductase, when partially inactivated by NADH and cyanide, was not altered as a substrate for the inactivating enzyme. The maize root inactivating enzyme was also shown to inactivate the nitrate reductase (NADH) in the pea leaf. It had no effect on the nitrate reductase from either Pseudomonas denitrificans or Nitrobacter agilis.
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PMID:Effects of a nitrate reductase inactivating enzyme and NAD(P)H on the nitrate reductase from higher plants and Neurospora. 23

The formation of hydrogen peroxide during the oxidation of NADH by purified preparations of cytochrome o has been demonstrated by employing three independent methods: polarographic, colorimetric, and fluorometric. The first two methods were used to assay for the accumulation of hydrogen peroxide and showed that hydrogen peroxide did accumulate as a product, but only about 30% of the oxygen consumed or 15 to 20% of the NADH oxidized was recoverable as hydrogen peroxide. This lack of 1:1 stoichiometry was not due to residual catalase activity in these preparations which could be eliminated by freeze-thawing. Thus, hydrogen peroxide may not be the sole or primary product of the NADH-cytochrome o oxidase reaction. The fluorometric assay could be coupled directly to the NADH-cytochrome o oxidase reaction in one medium, and this method showed that hydrogen peroxide was generated continuously from the beginning of the reaction in a 1:1 stoichiometry, hydrogen peroxide generated to NADH oxidized. This result suggests that hydrogen peroxide is an intermediate that can be trapped efficiently under the conditions of the fluorometric assay, whereas under the conditions of the first two assays most of the hydrogen peroxide generated undergoes further reaction. Exogenously added FAD or FMN increased the percentage of hydrogen peroxide that accumulated in the NADHcytochrome o oxidase reaction. Flavin is believed to act on the reductase side of cytochrome o so the increased percentage of hydrogen peroxide is not likely to result from the direct reaction of reduced flavin with oxygen.
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PMID:The formation of hydrogen peroxide during the oxidation of reduced nicotinamide adenine dinucleotide by cytochrome o from Vitreoscilla. 23 73

A NAD(P)H:flavin oxidoreductase, which produces FMNH2, one of the substrates for the luciferase reaction in bioluminescent bacteria, has been purified with the aid of affinity chromatography on epsilon-aminohexanoyl-FMN-Sepharose. The purified enzyme, isolated from Beneckea harveyi, had a specific activity of 89 mumol of NADH oxidized/min/mg of protein at 23 degrees in the presence of saturating FMN and NADH and appeared homogeneous by several criteria on polyacrylamide gel electrophoresis. A molecular weight of 24,000 was estimated both by gel filtration and and sodium dodecyl sulfate gel electrophoresis indicating that the enzyme is composed of a single polypeptide chain. Kinetic studies showed that the higher specificity of the enzyme for NADH than NADPH and for riboflavin and FMN than FAD was primarily due to variations in the Michaelis constants for the different substrates. Initial velocity studies with all pairs of substrates gave intersecting patterns supporting a sequential mechanism for the NAD(P)H:flavin oxidoreductase.
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PMID:Purification and properties of a NAD(P)H:flavin oxidoreductase from the luminous bacterium, Beneckea harveyi. 30 40

Using the powerful lachrymator (2-chlorobenzylidene)malononitrile as electron acceptor, two types of NAD(P)H dehydrogenases have been isolated from human blood. Crystallisation of the homogenous enzymes was performed in 50% polyethylene glycol solution. The enzymes (average molecular weight 18 000) are composed of only one polypeptide chain and have a very similar amino acid composition. B-side stereospecificity was determined with respect to the cofactor by gas chromatography-mass spectrometry for the reductase. Besides (2-chlorobenzylidene)malononitrile, 2,6-dichloroindophenol, methylene blue, 4-benzoquinone, FMN and FAD are also reduced using NADH or NADPH as hydrogen donor with the rates decreasing in the given order. Reduction of methemoglobin is observed only upon addition of methylene blue, FMN or FAD as carriers. (2-Chlorobenzylidene)malononitrile reduction is inhibited by most of the compounds known to be decouplers of oxidative phosphorylation.
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PMID:Benzylidenemalononitrile derivatives as substrates and inhibitors of a new NAD(P)H dehydrogenase of erythrocytes. Purification and crystallisation of two forms of the enzyme. 38 68

1. Ion-exchange chromatography resolves the methane mono-oxygenase from soluble extracts of Methylococcus capsulatus (Bath) into three fractions. 2. Fractions A and B are comparatively stable at 0 degrees C, whereas fraction C is very unstable unless kept in the presence of sodium thioglycollate (1-10 mM) or dithiothreitol (5-10mM). 3. The active component from fraction C was purified some 80-fold. 4. Purified component C has mol. wt. 42000. Its solutions are yellow with absorption maxima at 270 and 465 nm and a shoulder at 395 nm. The 465 nm peak is abolished by reduction with NADH or sodium dithionite, or by photoreduction in the presence of EDTA. A new spectral species, probably a neutral flavin semiquinone, is observed on partial reduction of component C. 5. No copper was detected in samples of purified component C, but the protein contains 1.3-1.5 atoms of iron/molecule. 6. On boiling, component C releases a yellow-green fluorescent material that has been identified as FAD from its absorption and fluorescence spectra and by t.l.c. 7. Component C contains 1 mol of FAD/mol of protein.
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PMID:Resolution of the methane mono-oxygenase of Methylococcus capsulatus (Bath) into three components. Purification and properties of component C, a flavoprotein. 41 77

2-Enoate-reductase, a previously unknown soluble enzyme is present in Clostridium kluyveri and another Clostridium species growing on (E)-2-butenoate. From the latter the reductase was purified 88-fold with an overall yield up to 74%. The enzyme was pure as judged by polyacrylamide gel electrophoresis with and without sodium dodecyl sulphate as well as by isoelectric focusing. The purification of the enzyme was performed in the presence of (E)-2-methyl-2-butenoate as substrate to keep the enzyme in the oxidized state and under anaerobic conditions. The purification procedure included an ammonium sulphate precipitation, chromatography on DEAE-Sepharose CL-6B, hydroxylapatite and Sepharose CL-6B. The enzyme reduces different alpha,beta-unsaturated carboxylate anions such as (E)-2-butenoate, (E)-2-methyl-2-butenoate, (E)-cinnamate and probably many others in a NADH-dependent reaction to the saturated carboxylate anions. Fumarate, 3-phenyl-2-propinate, 2-enoyl-methyl and CoA esters proved not to be substrates for the purified reductase. NADPH does not act as an electron donor. The enzyme was shown to have a molecular weight of about 450,000 by gel chromatography. It consists of subunits with a molecular weight of 78,000. Per subunit about 1 FAD, 3.5--3.8 atoms of iron and 4.0 labile sulphur atoms have been found indicating a conjugated iron-sulphur flavo-protein. Copper could not be detected. The isoelectric point was 8.4. As shown by absorption spectroscopy the enzyme can be reduced by NADH and reoxidized with dichloroindophenol, hexacyanoferrate III, oxygen and substrates. Addition of 8 mol p-hydroxymercuribenzoate to 1 mol subunit completely destroyed the activity of the reductase. So far no physiological role of the enzyme is known.
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PMID:Purification and some properties of a hitherto-unknown enzyme reducing the carbon-carbon double bond of alpha, beta-unsaturated carboxylate anions. 47 58

Assimilatory nitrate reductase (EC 1.6.6.1 NADH:nitrate oxidoreductase) from Chlorella vulgaris purified by affinity chromatography was found to be homogeneous as judged by electrophoresis on sodium dodecyl sulfate-polyacrylamide gel and by analytical ultracentrifugal techniques. The molecular weight of the intact enzyme and that of the enzyme dissociated in 6 M GuHCl, determined by sedimentation equilibrium studies, were 280,000 +/- 10,000 and 90,000 +/- 5,000, respectively. Comparable values were obtained using the S20,w value and the D20,w values in Svedberg's equation. The D20,w values were determined by laser light-scattering measurements. Active enzyme centrifugation showed that the monomer is an active species. A quantitative re-evaluation of the prosthetic groups present (FAD, heme, and molybdenum) was also made and was consistent with the conclusion that the active monomer contains three subunits as previously deduced by Solomonson et al. ((1975) J. Biol. Chem. 250, 4120). Electron micrographs showed images which corresponded to three subunits, supporting the data obtained by hydrodynamic studies. The enzyme is not cigar-shaped, as previously surmised, but has a roughly globular structure.
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PMID:Physical studies on assimilatory nitrate reductase from Chlorella vulgaris. 50 Jun 68

Spinach nitrate reductase complex previously inactivated by treatment with mercurials p-hydroxymercuribenzoate or p-hydroxymercuriphenyl sulphonate can be reactivated by incubation with dithioerythritol. The reactivation of NADH-diaphorase seems to be FAD-dependent, whereas that of FNH2-nitrate reductase is not. The requirement of FAD for NADH-inactivation of nitrate reductase treated with p-hydroxymercuribenzoate disappears after treatment with dithioerythritol.
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PMID:Nitrate reductase from Spinacea oleracea. FAD and the reactivation of the enzyme treated with p-Hydroxymercuribenzoate. 59 86

Cell extract from a strain of Propionibacterium acidi-propionici with high nitrate reductase (NaR) activity catalyzed nitrate reduction with glycerol phosphate, NADH, or lactate. The reaction was inhibited partially by fumarate or oxygen. NaR linked to methyl viologen was found mostly in particulate fractions. It was solubilized by treatment with Emulgen 810 and purified 46-fold by DEAE-cellulose, Sepharose 4B, and triple DEAE-Sephadex chromatographies in the presence of the detergent. It was rather labile but was stabilized by glycerol. The molecular weight was estimated to be 230,000 by Sepharose 4B gel filtration and the isoelectric point was pH 5.0-5.5. The pH optimum was at 6.5-7.5 and Km for nitrate was 0.1 mM. As electron donors, methyl and benzyl viologen were utilized well but FAD and FMN were fairly ineffective. Chlorate was an active acceptor as well as nitrate. Azide, cyanide, and thiocyanate inhibited NaR. On adding 1 mM tungstate to the growing medium, the NaR level in grown cells was lowered; addition of 0.01 mM molybdate restored the activity partially. NaR is suggested to be a molybdo-protein, similar to this enzyme from other bacteria.
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PMID:A study on nitrate reductase from Propionibacterium acidi-propionici. 62 3

Formate dehydrogenase (EC 1.2.1.2) from Pseudomonas oxalaticus has been isolated and characterized. The enzyme (molecular weight 315000) is a complex flavoprotein containing 2 FMN, 18--25 non-heme iron atoms and 15--20 acid-labile sulphides. In the last step of the purification, a sucrose gradient centrifugation, a second catalytically active species has been found apparently originating from a dissociation of the enzyme into two equal subunits. The enzyme is specific toward its natural substrate formate. It transfers electrons to NAD+, oxygen, ferricyanide, and a lot of nonphysiological acceptors (dyes). In addition electrons are transferred from NADH to these acceptors. The (reversible) removal of FMN requires a reduction step. Reincorporation has been followed by the reappearance of the reactivity against formate and by fluorescence titration. The deflavo enzyme also binds FAD and riboflavin. The resulting enzyme species show characteristic catalytic abilities. Activity against formate is peculiar to the FMN species.
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PMID:Formate dehydrogenase from Pseudomonas oxalaticus. 63 Nov 30


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