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)

Cyclohexylamine oxidase was purified 90-fold from cell-free extracts of Pseudomonas sp. capable of assimilating sodium cyclamate. The purified enzyme was homogeneous in disc electrophoresis, and the molecular weight was found to be approximately 80,000 by gel filtration. The enzyme catalyzed the following reaction: cyclohexylamine+O2+H2O leads to cyclohexanone+NH3+H2O2. The enzyme thus can be classified as an amine oxidase; it utilized oxygen as the ultimate electron acceptor. The pH optimum of the reaction was 6.8 and the apparent Km value for cyclohexylamine was 2.5 X 10(-4) M. The enzyme was highly specific for the deamination of alicyclic primary amines such as cyclohexylamine, but was found to be inactive toward ordinary amines used as substrates for amine oxidases. The enzyme solution was yellow in color and showed a typical flavoprotein spectrum; the addition of cyclohexylamine under anaerobic conditions caused reduction of the flavin in the native enzyme. The flavin of the prosthetic group was identified as FAD by thin layer chromatography. The participation of sulfhydryl groups in the enzymic action was also suggested by the observation that the enzyme activity was inhibited in the presence of PCMB and could be recovered by the addition of glutathione.
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PMID:Purification and some properties of cyclohexylamine oxidase from a Pseudomonas sp. 1 51

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

1. The presence of xanthine was required for the inhibition of bovine milk xanthine oxidase by o-iodosobenzoate, iodoacetamide, hydrogen peroxide or p-chloromercuribenzoate. 2. Inactivation by p-chloromercuribenzoate was very rapid, was reversed by cysteine and was less in the presence of FAD. Lineweaver-Burk plots showed that the inactivation by p-chloromercuribenzoate was competitive with substrate. 3. Inactivation by o-iodosobenzoate, iodoacetamide or hydrogen peroxide could not be reversed by cysteine or xanthine. However, the presence of xanthine during the incubation with inhibitor protected the enzyme against o-iodosobenzoate but not against iodoacetamide or hydrogen peroxide. 4. p-Chloromercuribenzoate protected the enzyme against inactivation by hydrogen peroxide.
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PMID:Xanthine oxidase inactivation by reagents that modify thiol groups. 562 93

The total -SH content of purified NADPH-cytochrome P-450 reductase (NADPH: ferricytochrome oxidoreductase, EC 1.6.2.4) from rabbit liver microsomes accessible to an excess equivalent of PCMB was 7.0 +/- 0.3 mol thiol groups/mol protein. The modification of four -SH groups at low concentration of PCMB stimulated the activity of the enzyme. On the other hand, further blocking of -SH groups (6-7 mol -SH groups/Mol protein) with an excess amount of PCMB completely inhibited cytochrome c (or DCPI) reductase activity. The fluorescence quenching of the flavin was rapidly removed by binding of PCMB to a fifth and sixth -SH group during a gradual titration. Kinetic and fluorimetric analyses confirmed the suggestion that these two -SH groups essential for catalytic function were partly protected by NADP+ or 2'-AMP against the reaction with PCMB. Excess PCMB begins to compete with the ligand preincubated with the enzyme. The spectral perturbation on the addition of approx. 6-7 equiv. PCMB/mol enzyme is accompanied by a slight blue shift of the absorbance maximum at 380 nm, with the appearance of a pronounced shoulder at 475 nm. In contrast to the native enzyme, 3-electron-reduced semiquinone form of PCMB-treated enzyme showed the same absorption spectrum as 1-electron-reduced semiquinone which has an absorption maximum at 585 nm with a broad shoulder around 635 nm. An inhibitory effect may be attributable to the fact that NADPH is less accessible to the FAD binding site as well as the pyridine nucleotide binding site, since the rate of FAD reduction becomes extremely slow after complete modification.
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PMID:Location of functional -SH groups in NADPH-cytochrome P-450 reductase from rabbit liver microsomes. 679 74

NADPH-dependent 2,6-dichlorophenol-indophenol (DCIP) reductase activity in the homogenate of phagocytosing pig polymorphonuclear leucocytes was twice that of the resting cells and the activity in the phagocytic vesicles corresponded to the activity increment due to phagocytosis. The apparent Km value of the reductase activity in the vesicles for NADPH was 30 microM, which is similar to that of the NADPH-dependent superoxide (O2-) formation. Increasing the DCIP reductase activity by increasing the DCIP concentration caused a decrease in the O2- -forming activity, the NADPH oxidation rate being constant and independent of the dye concentration. p-Chloromercuribenzoate and cetyltrimethylammonium bromide at low concentrations inhibited the O2- -forming activity of the vesicles without inhibiting the DCIP reductase. Quinacrine inhibited both O2- formation and DCIP reduction. The DCIP reductase activity could be extracted with a mixture of deoxycholate and Tween-20, which extracts the O2- -forming activity. The reductase activity in the extract was enhanced 2-fold by the addition of FAD, and its apparent Km was 0.085 microM. These results indicate that the NADPH-dependent DCIP reductase activity of the phagocytic vesicles is catalysed by a flavin-containing component of the O2- -forming system.
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PMID:NADPH-dependent reduction of 2,6-dichlorophenol-indophenol by the phagocytic vesicles of pig polymorphonuclear leucocytes. 686 Mar 11

Effectors and products of enzymatic diiodotyrosine (DIT) deiodination by a cytosolic fraction of pig liver hab been investigated. 13% of the degraded 131I-DIT was found as monoiodotyrosine by thin layer chromatography. The main quantity of the deiodinated DIT was found on the start point of the chromatogram bound to enzyme protein. Tyrosine as a reaction product of enzymatic deiodination of [14C]-IT could not be identified exactly. The liver cytosolic deiodinase is activated by pyruvate; the extent of activation depends on th pyruvate concentration. Diiodohydroxyphenylpyruvate as a product of transamination and theoretically possible intermediate product could be excluded. NADPH 2 and sodium dithionite activated the deiodinase to 1/3, sodium dithionite together with FAD to 1/2 the amount of which was determined for the action of pyruvate. The enzymatic activity in the presence of pyruvate and NADPH2, respectively NADPH2/FAD is identical with the sum of the single activities. The effect of dithionite and sulfite on deiodinase activity depends on the concentration: low effector concentrations increase, while high concentrations decrease the enzyme activity. The liver plasma deiodinase was inactivate quantitatively by reaction with 10(-4) M PCMB; by reaction with 10(-4) M DTNB or NEM the inactivation was 40% only. The inactivation of deiodinase by PCMB was quantitative reversible by cysteine, while inactivation by DTNB was reversible by cysteine to maximal 70% only. Differences between cytosolic and microsomal deiodinases are discussed also in regard to the mechanism of DIT-deiodination by a liver cytosolic fraction with direct participation of SH-groups.
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PMID:[Effectors and products of enzymatic diiodotyrosine deiodination by a plasmatic fraction from pig liver]. 730 42

Monodehydroascorbate radical (MDA) reductase, an FAD-enzyme, is the first enzyme to be identified whose substrate is an organic radical and catalyzes the reduction of MDA to ascorbate by NAD(P)H. Its cDNA has been cloned from cucumber seedlings (Sano, S., and Asada, K. (1994) Plant Cell Physiol. 35, 425-437), and a plasmid was constructed in the present study that allowed a high level expression in Escherichia coli of the cDNA-encoding MDA reductase using the T7 RNA polymerase expression system. The recombinant MDA reductase was purified to a crystalline state, with a yield of over 20 mg/liter of culture, and it exhibited spectroscopic properties of the FAD similar to those of the enzyme purified from cucumber fruits during redox reactions with NADH and MDA. The red semiquinone of the FAD of MDA reductase was generated by photoreduction. p-Chloromercuribenzoate inhibited the reduction of the enzyme-FAD by NADH, and dicumarol suppressed electron transfer from the reduced enzyme to MDA. The specificity of electron acceptors of the recombinant enzyme appeared to be similar to that of MDA reductase, even though the amino acid sequence encoded by the cDNA was somewhat different from that of the enzyme purified from cucumber fruits. The Km values for NADH and NADPH of the recombinant enzyme indicated a high affinity of the enzyme for NADH. The reaction catalyzed by the enzyme did not exhibit saturation kinetics with MDA up to 3 microM. A second order rate constant for the reduction of the enzyme-FAD with NADH was 1.25 x 10(8) M-1 s-1, as determined by a stopped-flow method, and its value decreased with increases in ionic strength, an indication of the enhanced electrostatic guidance of NADH to the enzyme-FAD.
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PMID:Molecular characterization of monodehydroascorbate radical reductase from cucumber highly expressed in Escherichia coli. 754 69