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)

A phagocytic vesicle fraction with high NADPH-dependent superoxide-forming activity was obtained in large quantity from pig blood polymorphonuclear leucocytes, phagocytosing oil droplets in the presence of cyanide. The activity of the homogenate of the phagocytosing cells was 40 times that of the resting cells, and 70% of the activity in the homogenate was recovered in the phagocytic vesicle fraction. Essentially all of the superoxide-forming activity was extracted by repeated extraction with a mixture containing deoxycholate and Tween 20. The extract had a superoxide-forming activity of 1 mumol/min per mg of protein with NADPH, and one-fifth of this with NADH, Km values being similar to those of the vesicle fraction (40 microM for NADPH and 400 microM for NADH). A stoichiometric relationship of 1:2 for NADPH oxidation and superoxide formation was obtained, in agreement with the reaction NADPH +2O2 leads to NADP+ + 2O2 -. + H+. The activity of the extract was enhanced 2-fold by the addition of FAD, suggesting that the flavin is a component of the enzyme system. The Km value for FAD was 0.077 microM. The activities in both vesicle fraction and extract were labile even on refrigeration, but could be kept for several months at -70 degrees C.
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PMID:Superoxide-forming NADPH oxidase preparation of pig polymorphonuclear leucocyte. 629 59

By means of a cytochemical technique, hydrogen peroxide formation was located on the endothelial cell surface (predominantly the luminal aspect) of capillaries obtained by collagenase digestion of rat thyroid. The cyanide-insensitive H2O2 formation required aerobic conditions and NAD(P)H as substrate. FAD could also stimulate the reaction, but not xanthine. The cytochemical reaction was blocked by a non-penetrating protein inhibitor. The observations are interpreted as evidence of a plasmalemma-bound H2O2-generating enzyme. The findings indicate that microvascular endothelial cells are involved in the release of activated oxygen species, which might have important pathophysiologic implications.
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PMID:Cytochemical demonstration of an enzymatic production of hydrogen peroxide on the surface of isolated thyroid capillaries. 629 36

The spectral properties of a particulate fraction of human polymorphonuclear neutrophils capable of oxidizing NADPH were studied before and after depletion of myeloperoxidase by KCl treatment. Difference spectra (dithionite reduced minus oxidized) at 77 K of non-extracted particles showed peaks of a b-type cytochrome at 556, 527 and 425 nm and of myeloperoxidase at 636 and 474 nm. Extraction of myeloperoxidase led to a 4-5-fold increase in the size of the cytochrome b peaks. In non-extracted particles, the CO-reduced spectra at 77 K revealed a typical CO-reduced myeloperoxidase complex with new peaks at 625-630 and 462 nm, and a limited shift of the Soret band of reduced cytochrome b from 425 to 424-423 nm. The same shift was observed for cytochrome b in extracted particles. Photoirradiation of the CO-dithionite-reduced particles resulted in a back shift of the CO-reduced peaks to their original positions in the reduced spectrum. Concomitantly, the size of the peaks both for myeloperoxidase and cytochrome b was increased, indicating photoreduction. Cytochrome b and myeloperoxidase in neutrophil particles were poorly reduced by NADPH; reduction occurred upon photoirradiation. FAD and FMN added to particles in the presence of NADPH were photoreduced concomitantly with cytochrome b. Addition of phorbol myristate acetate to intact neutrophils in the presence of glucose resulted in CO- and cyanide-insensitive respiration, accumulation of O-2, and also in reduction of cytochrome b. The lag required to reach the steady-state production of O-2 was equal to the lag required for cytochrome b to reach a plateau of reduction. The data are consistent with the idea that cytochrome b in neutrophils might belong to a branched pathway that is not rate-limiting in the cyanide-resistant respiration of the neutrophils.
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PMID:Examination of the oxidase function of the b-type cytochrome in human polymorphonuclear leucocytes. 632 Aug 72

A highly specific inducible membrane-bound 4-pyridoxic acid dehydrogenase has been solubilized and purified to apparent homogeneity from Pseudomonas MA-1 grown with pyridoxine as a sole source of carbon and nitrogen. The undenatured enzyme migrates as a single band on gel electrophoresis; denatured preparations show two barely resolved bands (Mr = 63,000 and 61,000). Undenatured preparations aggregate readily, as evidenced by Mr values of 148,000, 470,000, and greater than 670,000 obtained by density gradient centrifugation or by gel filtration under various conditions. The enzyme contains FAD but no Fe or acid-labile S; an average minimum molecular weight of 131,000 was calculated from the FAD content. In the presence of 2,6-dichloroindophenol, the enzyme dehydrogenates 4-pyridoxic acid to the corresponding aldehyde; this reaction is not inhibited by CN-. At the pH optimum of 8.0, a Vm of approximately 7.0 mumol min-1 mg-1 and a Km of 9 microM were obtained. 2,6-Dichloroindophenol, phenazine methosulfate, and menadione are effective electron acceptors; ubiquinones are less active, while NAD, FAD, and O2 are inactive. However, in membrane fractions, oxygen supports 4-pyridoxic acid oxidation via a CN--sensitive electron transport chain, indicating that the dehydrogenase probably is coupled to ATP generation in such preparations.
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PMID:The bacterial oxidation of vitamin B6. 4-Pyridoxic acid dehydrogenase: a membrane-bound enzyme from Pseudomonas MA-1. 634 42

Purine hydroxylase II from Aspergillus nidulans has been purified to near homogeneity. The enzyme has a pI of 5.7, a molecular weight of 300,000, and two subunits with molecular weight of 153,000 each. The enzyme contains 2 FAD, 2 molybdenum atoms, and 4 (2 Fe-2S) iron-sulfur centers per molecule and exhibits broad specificity for reducing and oxidizing substrates. Among the more notable characteristics are the ability to oxidize hypoxanthine and nicotinic acid but not xanthine and virtually complete inactivity with oxygen. Moreover, while the enzyme is inactivated by borate and methanol, it is very resistant to cyanide and arsenite and it not inactivated by allopurinol. At infinite concentrations of reducing and oxidizing substrates, the Km for hypoxanthine was 119 microM, for nicotinic acid was 136 microM, and for NAD+ was 525 microM.
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PMID:Purification and properties of purine hydroxylase II from Aspergillus nidulans. 636 61

The properties and requirements of squalene epoxidase and effects of some inhibitors were investigated in the pathogenic yeast Candida albicans. A washed 'microsomal' fraction converted radiolabelled squalene to 2,3-oxidosqualene and lanosterol. Minimum requirements for activity were molecular oxygen, NADH or NADPH, and FAD. Epoxidase activity was stimulated by up to 100% by addition of the soluble cytoplasmic fraction, which itself contained negligible epoxidase activity. This stimulation was most powerful at low concentrations of enzyme, or high concentrations of squalene. Divalent cations did not stimulate activity and EDTA was not inhibitory. An apparent Km for squalene of 50 microM was determined in the presence of soluble cytoplasm. Epoxidase activity was destroyed by Triton X-100, deoxycholate or Cu2+, and partially inhibited by thiol reagents, rotenone and antimycin A. The enzyme was not inhibited by cyanide or by several inhibitors of cytochrome P-450.
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PMID:Properties of a particulate squalene epoxidase from Candida albicans. 637 56

The molecular basis for the action of two natural inactivator proteins, isolated from rice and corn, on a purified assimilatory nitrate reductase has been examined by several physical techniques. Incubation of purified Chlorella nitrate reductase with either rice inactivator protein or corn inactivator protein results in a loss of NADH:nitrate reductase and the associated partial activity, NADH:cytochrome c reductase, but no loss in nitrate-reducing activity with reduced methyl viologen as the electron donor. The molecular weight of the reduced methyl viologen:nitrate reductase species, determined by sedimentation equilibrium in the Beckman airfuge after complete inactivation with rice inactivator protein or with corn inactivator protein, was 595,000 and 283,000, respectively, compared to a molecular weight of 376,000 for the untreated control determined under the same conditions. Two protein peaks were observed after molecular-sieve chromatography on Sephacryl S-300 of nitrate reductase inactivated by corn inactivator protein. The Stokes radii of these fragments were 68 and 24 A, compared to a value of 81 A for untreated nitrate reductase. The large fragment contained molybdenum and heme but no flavin, and had nitrate-reducing activity with reduced methyl viologen as electron donor. The small fragment contained FAD but had no NADH:cytochrome c reductase or nitrate-reducing activities. Molecular weights determined by sodium dodecyl sulfate-gel electrophoresis were 67,000 and 28,000 for the large and small fragments, respectively, compared to a subunit molecular weight of 99,000 determined for the untreated control. No change in subunit molecular weight of nitrate reductase after inactivation by rice inactivator protein was observed. These results indicate that rice inactivator protein acts by binding to nitrate reductase. The stoichiometry of binding is 1-2 molecules of rice inactivator protein to one tetrameric molecule of nitrate reductase. Corn inactivator protein, in contrast, acts by cleavage of a Mr 30,000 fragment from nitrate reductase which is associated with FAD. The remaining fragment is a tetramer of Mr 70,000 subunits which retains nitrate-reducing activity and contains molybdenum and heme but has no NADH:dehydrogenase activity. The action of rice inactivator protein was partially prevented by NADH and completely prevented by a combination of NADH and cyanide, while the action of corn inactivator protein was not significantly affected by these effectors.
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PMID:Mode of action of natural inactivator proteins from corn and rice on a purified assimilatory nitrate reductase. 654 59

By techniques involving differential centrifugation and specific precipitation with CaCl2, it was shown that dimethylamine and trimethylamine mono-oxygenase activities co-sediment with NADPH-cytochrome c reductase activity in sphaeroplast lysates of Candida utilis grown on trimethylamine as sole nitrogen source. Since the active fraction also contained low levels of cytochromes P-450 and P-420, it was concluded that the two amine mono-oxygenases are located in the smooth endoplasmic reticulum and thus end up in the microsomal fraction on cell fractionation. Ten to twenty-fold enrichment of mono-oxygenase specific activity could be achieved by separation of activity from soluble protein by centrifugation or gel filtration. Cell-free extracts prepared in the absence of FAD showed only very low mono-oxygenase activity for either substrate. Some activity could be restored by addition of flavin nucleotides: there was a fivefold stimulation by FAD and a fourfold stimulation by FMN. All trimethylamine mono-oxygenase activity was lost when a partially purified preparation containing both activities was incubated for more than 24 h at 0 degrees C, suggesting that separate enzymes are responsible for the oxidation of secondary and tertiary amines. The enzyme preparation oxidized a wide range of secondary alkylamines up to dibutylamine and tertiary alkylamines up to tributylamine. Primary amines, choline, di- and triethanolamine, spermine, spermidine and substituted anilines were not oxidized. NADH had a lower apparent Km value and higher Vmax value than NADPH. Secondary and tertiary alkylamines containing more than one kind of alkyl group gave more than one kind of aldehyde on oxidation. Stoicheiometry determinations showed a consumption of 1 mol NAD(P)H and 1 mol O2 per mol aldehyde formed. Carbon monoxide, cyanide, proadifen hydrochloride (SKF 525-A), mercurials and mercaptoethanol all inhibited both activities.
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PMID:Subcellular localization and properties of partially purified dimethylamine and trimethylamine mono-oxygenase activities in Candida utilis. 654 83

Assimilatory nitrate reductase (NAD(P)H-nitrate oxidoreductase, EC 1.6.6.2) from the green alga Ankistrodesmus braunii can be purified to homogeneity by dye-ligand chromatography on blue-Sepharose. The purified enzyme, whose turnover number is 623 s-1, presents an optimum pH of 7.5 and Km values of 13 microM, 23 microM and 0.15 mM for NADH, NADPH and nitrate, respectively. The NADH-nitrate reductase activity exhibits an iso ping pong bi bi kinetic mechanism. The molecular weight of the native nitrate reductase is 467 400, while that of its subunits is 58 750. These values suggest an octameric structure for the enzyme, which has been confirmed by electron microscopy. As deduced from spectrophotometric and fluorimetric studies, the enzyme contains FAD and cytochrome b-557 as prosthetic groups. FAD is not covalently bound to the protein and is easily dissociated in diluted solutions from the enzyme. Its apparent Km value is 4 nM, indicative of a high affinity of the enzyme for FAD. The results of the quantitative analyses of prosthetic groups indicate that nitrate reductase contains four molecules of flavin, four heme irons, and two atoms of molybdenum. The three components act sequentially transferring electrons from reduced pyridine nucleotides to nitrate, thus forming a short electron transport chain along the protein. A mechanism is proposed for the redox interconversion of the nitrate reductase activity. Inactivation seems to occur by formation of a stable complex of reduced enzyme with cyanide or superoxide, while reactivation is a consequence of reoxidation of the inactive enzyme. Both reactions imply the transfer of only one electron.
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PMID:Assimilatory nitrate reductase from the green alga Ankistrodesmus braunii. 668 79

Whole cells of Candida boidinii grown on di- or tri-methylamine as sole nitrogen source readily oxidized both amines. The oxidation was potently inhibited by carbon monoxide. Cell-free extracts required the presence of 20 microM FAD before mono-oxygenase activity with both amines could be demonstrated. NADH was a better electron donor than NADPH. Activity was present in cells grown on secondary and tertiary amines but not on primary amines, and was detected in a number of different yeasts. Enzyme activity could be sedimented at 187 000 x g, and was associated with NADPH-cytochrome c reductase activity. It is thus probably microsomal. Activity was inhibited by cyanide, mercaptoethanol, carbon monoxide and proadifen hydrochloride (SKF 525-A).
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PMID:Oxidation of dimethylamine and trimethylamine in methazotrophic yeasts by microsomal mono-oxygenases sensitive to carbon monoxide. 687 Sep 1


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