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

1. A continuous spectrophotometric determination of rat hepatic microsomal anaerobic azo reductase activity has been developed. 2. The addition of soluble flavins (riboflavin, FMN or FAD) greatly increased this NADPH-dependent activity towards a number of azo substrates. 3. Investigations with amaranth as substrate gave an apparent Km of 34 microM and Vmax. of 4 nmol/min per mg of microsomal protein. The inclusion of a fixed concentration of FMN increased Vmax. and greatly decreased Km, the magnitude of these changes reflecting the concentration of flavin present. 4. Investigations using a fixed amaranth concentration over a range of flavin concentrations gave biphasic double-reciprocal plots with two apparent Km and Vmax. values. 5. Pretreatment of animals with cobaltous chloride, 2-allyl-2-isopropylacetamide, carbon tetrachloride, phenobarbitone and 3-methylcholanthrene altered azo reductase activity in parallel with changes in cytochrome P-450 content. 6. The significance of these results is discussed in terms of the electron-transfer components present in the hepatic microsomal fraction.
 ...
PMID:A continuous spectrophotometric determination of hepatic microsomal azo reductase activity and its dependence on cytochrome P-450. 709 14

Norcocaine nitroxide was found to be produced via the one-electron oxidation of N-hydroxynorcocaine by hepatic microsomal enzymes from induced and noninduced rats, hamsters, and mice in the presence of an NADPH-generating system. This reaction was demonstrated to be mediated by cytochrome P-450 as suggested by induction experiments using phenobarbital, which markedly enhanced the production of this nitroxide, and by the inhibition of this monooxygenase by metyrapone, which depressed the formation of this free radical. Unlike other nitroxides, norcocaine nitroxide was rapidly reduced by flavoproteins such as cytochrome P-450 reductase and FAD-monooxygenase, but not cytochrome P-450. We believe that since NADPH is consumed during the futile cycling of N-hydroxynorcocaine/norcocaine nitroxide and since NADPH is an essential cofactor of the glutathione reductase system, diminished reduced nucleotide may lead to depressed levels of cellular glutathione. In this manner, we theorize that cocaine initiates hepatotoxicity.
 ...
PMID:Norcocaine nitroxide. A potential hepatotoxic metabolite of cocaine. 709 46

Sex-related differences in the activity of hepatic FAD-containing monooxygenase (FAD-M) were found in C3H/St mice. Adult female mice had enzyme activities nearly two-fold greater than male mice and these differences, which were absent in sexually immature mice, became apparent at the onset of puberty. The sex differences in hepatic FAD-M appeared to be mediated through the suppressive effect of testosterone; castration of male mice enhanced enzyme activity, while androgenic replacement returned activities to control levels. Testosterone's suppressive effect was found to be relatively specific for hepatic FAD-M. Treatment of castrated male mice with both the anti-androgen flutamide and testosterone returned enzyme activity to control levels, suggesting that testosterone's regulation of hepatic microsomal FAD-M is receptor-mediated. Female gonadectomy had no effect on this enzyme's activity.
 ...
PMID:Androgenic suppression of mouse hepatic FAD-containing monooxygenase activity. 713 54

The addition of substrates to microsomal FAD-containing-monooxygenase (EC 1.14.13.8) is an ordered process in which FAD reduction by NADPH is necessarily the first step. This reduction by NADPH is markedly biphasic, and when analyzed for two phases, rate constants of 2.46 s-1 and 0.88 s-1 are obtained at 4 degrees C, pH 7.2. The Kd (8 microM) for the binding of NADPH to the flavoprotein prior to reduction demonstrates a tight binding. Reduction by NADH also occurs and is similar in all respects to that observed with NADPH with the exception that the Kd is larger (0.167 mM) and easily measured. Oxygenatable substrates pre-equilibrated with the enzyme do not change the reduction in any way. NADP+ equilibrated with the enzyme prior to reduction removes the faster phase of the reaction leaving only the slower 0.88 s-1 phase. However, NADP+ rapidly mixed with the enzyme at the time of reduction does not affect the biphasic nature of the reduction, indicating that the binding of NADP+ to the enzyme results in a comparatively slow change of the form of the enzyme responsible for the fast phase into that which exhibits the slower rate. A primary deuterium isotope effect of approximately 6-fold has been observed on both phases of the reduction using (4R)-[4-2H]NADPH. This is strong evidence that both phases are due to primary reduction steps and that the enzyme preferentially abstracts the (4R)-proton. n-Octylamine, frequently present in turnover reactions because of its activating effect on the overall turnover rate of the enzyme, has a slightly inhibitory effect on the reduction step. We also show steady state kinetic patterns determined at both 25 degrees C and 15 degrees C which substantiate the results of Poulsen and Ziegler (J. Biol. Chem. 254, 6449-6455) which were determined at 37 degrees C.
 ...
PMID:The reductive half-reaction of liver microsomal FAD-containing monooxygenase. 721 2

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.
 ...
PMID:[Effectors and products of enzymatic diiodotyrosine deiodination by a plasmatic fraction from pig liver]. 730 42

Stopped flow kinetic studies have been used to demonstrate three features of the enzymatic mechanism of the microsomal FAD-containing monooxygenase from hog liver. First, in contrast to the bacterial flavin-containing monooxygenases, reduction of the FAD is independent of substrate. Second, the rate of the reaction of reduced enzyme with oxygen to form the C(4a)-peroxyflavin intermediate is independent of substrate. Third, the rate of transformation of the C(4a)-peroxyflavin to oxidized FAD is substrate-dependent. These results are in agreement with the mechanism, determined by steady state kinetic studies (Poulsen, L.L., and Ziegler, D.M. (1979) J. Biol. Chem. 254, 6449-6455), which predicts that the reduced flavin reacts with oxygen before combination with substrate.
 ...
PMID:Transient kinetic study of liver microsomal FAD-containing monooxygenase. 737 47

We have characterized a chemically reactive propranolol (PL) metabolite which binds to proteins in rat liver microsomes. During incubation with rat liver microsomes (1 mg of protein) fortified with an NADPH-generating system, 4-hydroxypropranolol (4-OH-PL) quickly disappeared from the reaction medium, but none of the possible metabolite peaks was detected under the high-performance liquid chromatographic conditions used. The consumption of 4-OH-PL depended on microsomes and NADPH. The reaction was not affected by inhibitors of cytochrome P450 or FAD monooxygenase, but was markedly diminished in the presence of cytosol and ascorbic acid. The effect of cytosol was inhibited by potassium cyanide but not by sodium benzoate or dimethyl sulfoxide, and was also not affected by heating at 60 degrees C for 30 min, suggesting that superoxide (SO) ion was involved in the reaction and that it was blocked by superoxide dismutase (SOD) present in the cytosol. Cu,Zn-SOD, purified from cytosol, effectively mimicked the suppressive effect of cytosol. Incubation of 4-OH-PL in an SO-generating system of xanthine and xanthine oxidase generated 1,4-naphthoquinone (1,4-NQ), which was identified by TLC, HPLC, and GC/MS. 1,4-NQ was also formed in microsomal incubates containing NADPH and small amounts of microsomes (below 0.1 mg of protein). These results indicate that 4-OH-PL is converted by SO, or some reactive oxygen species derived from it, to 1,4-NQ which binds to proteins and is one of the reactive metabolites of PL.
 ...
PMID:Characterization of a chemically reactive propranolol metabolite that binds to microsomal proteins of rat liver. 754 55

NADPH-cytochrome P-450 oxidoreductase (EC 1.6.2.4) was purified from the microsomal fraction of tobacco (Nicotiana tabacum) BY2 cells by chromatography on two anion-exchange columns and 2',5' ADP-Sepharose 4B column. The purified enzyme showed a single protein band with a molecular weight of 79 kDa on SDS-PAGE and exhibited a typical flavoprotein redox spectrum, indicating the presence of an equimolar quantity of FAD and FMN. This enzyme followed Michaelis-Menten Kinetics with Km values of 24 microM for NADPH and 16 microM for cytochrome c. An in vitro reconstituted system of the purified reductase with a partially purified tobacco cytochrome P-450 preparation showed the cinnamic acid 4-hydroxylase activity at the rate of 14 pmol min-1 nmol-1 P-450 protein and with a purified rabbit P-4502C14 catalyzed N-demethylation of aminopyrine at the rate of 6 pmol min-1 nmol-1 P-450 protein. Polyclonal antibodies raised against the purified reductase reacted with tobacco reductase but not with yeast reductase on Western blot analysis. Anti-yeast reductase antibodies did not react with the tobacco reductase. This result indicate that the tobacco reductase was immunochemically different from the yeast reductase. The anti-tobacco reductase antibodies totally inhibited the tobacco reductase activity, but not the yeast reductase. Also, Western blot analyses using the anti-tobacco reductase antibodies revealed that leaves, roots and shoots of Nicotiana tabacum plants contained an equal amount of the reductase protein. From these results, it was suggested that there are different antibody binding sites, which certainly participate in enzyme activity, between tobacco and yeast reductase.
 ...
PMID:Purification and immunochemical characteristics of NADPH-cytochrome P-450 oxidoreductase from tobacco cultured cells. 781 31

Fatty acid subterminal (omega-1 approximately omega-3) hydroxylase of the fungus Fusarium oxysporum was solubilized from the microsomal fraction and partially purified. The hydroxylase activity was recovered into a single active fraction, and its spectral nature showed the presence of cytochrome P-450 (P-450). Fatty acid hydroxylase activity was markedly restored upon addition of FAD, FMN, and/or hemin to the eluted fraction. The fraction also exhibited other properties characteristic of both a hemeprotein and a flavin-containing reductase. These results are highly indicative that the fungal hydroxylase is a fused protein containing both P-450 and its reductase domains. In this aspect the fungal enzyme resembles bacterial P-450BM3, although it is membrane-bound unlike the bacterial counterpart.
 ...
PMID:Fatty acid hydroxylase of the fungus Fusarium oxysporum is possibly a fused protein of cytochrome P-450 and its reductase. 803 65

Neutrophil-membrane-associated NADPH-cytochrome c reductase and cytochrome b558 were separately eluted and highly purified by a combination of ion-exchange Sepharose, N-amino-octylagarose, 2',5'-ADP-Sepharose and heparin-Sepharose column chromatographies. The purified cytochrome c reductase with an apparent molecular mass of 68 kDa contained FMN and FAD (FMN/FAD approx. 1). Cytochrome b558 prepared in the presence of phospholipids and FAD showed marked O2-.-producing activity (Vmax., 8.53 mumol of O2-./min per mg of cytochrome; Km for NADPH 58.8 microM) in a cell-free assay system consisting of cytosol, arachidonate and GTP[S]. However, when it was obtained without FAD added to the purification process, it had negligible FAD and little or no O2-.-forming activity in the reconstituted system. The NADPH oxidase activity was not markedly stimulated on incubation of the purified reductase with either flavinated or flavin-depleted cytochrome b558 in the cell-free system, suggesting that the reductase is not likely to be involved in neutrophil O2-. generation. The purified reductase cross-reacted with polyclonal antibodies against both hepatic NADPH-cytochrome P-450 reductase and a synthetic peptide, ILVGPGTGIAPFRSF, which indicates residues 529-543 located in the glycine-rich NADPH-binding domain of the P-450 reductase, but cytochrome b558 did not produce any immunoreactive bands to these antibodies. These antibodies also produced a positive reaction with a 76 kDa protein from dimethyl sulphoxide-induced HL-60-cell microsomes. After solubilization of the microsomal membranes, the 76 kDa protein was readily converted into a partially proteolysed form (68 kDa) even in the presence of antiproteases. In addition, the microsomal fraction shows a CO difference spectrum with a peak at about 454 nm and a trough at 476 nm in the presence of dithionite, indicating the presence of a cytochrome P-450-like haemoprotein.
 ...
PMID:NADPH-cytochrome c reductase from human neutrophil membranes: purification, characterization and localization. 811 Jan 98


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>