Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

The inhibition of rabbit skeletal muscle glycogen phosphorylase b by FAD and its analogues with substitutes in the position 8 has been studied. The value of half-saturation, [I]0,5, for inhibitors increases in the following order: FAD (44 microM), 8 alpha-hydroxy-FAD (60 microM), 8-dimethylamino (nor)-FAD (69 microM), 8 alpha-(N-acetyl-L-cystein-S-yl)-FAD (106 microM). From the comparison of these values with those obtained earlier for FMN analogues, it follows that in the case of FAD the half-saturation value is less sensitive to modification of the position 8 in the flavin isoalloxazine ring. The existence of the glycogen phosphorylase b FAD-complex has been proved by the spectrophotometry and sedimentation methods. The positions of maxima of optical absorption of the enzyme-bound FAD in the 300-500 nm region are identical with corresponding positions for FMN. FAD has been shown to hinder the AMP-induced transition of dimeric form of the enzyme to tetrameric one.
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PMID:[Interaction of muscle glycogen phosphorylase B with flavin-adenine dinucleotide and its analogs]. 392 80

Medium-chain and long-chain acyl-CoA dehydrogenases from rat liver have been purified in two forms, holoenzymes containing FAD and apoenzymes which do not contain this cofactor. In contrast, short-chain acyl-CoA dehydrogenase can only be isolated as the holoenzyme. Marked differences in the reactivity to organic sulfhydryl reagents were observed between the apo and holo forms of these enzymes. While the two apoenzymes were severely inactivated by N-ethylmaleimide (NEM), p-chloromercuribenzoate (pCMB), and iodoacetate (IAA), the two corresponding holoenzymes were not susceptible to these reagents. The inactivation of the two apoenzymes by NEM followed pseudo-first order kinetics. Incubation of the apoenzymes with FAD completely prevented the inactivation by the organic sulfhydryl reagents. Methylmercury halides (iodide or chloride) inactivated both the apo and holo forms of medium-chain and long-chain acyl-CoA dehydrogenases. On the other hand, holo-short-chain acyl-CoA dehydrogenase behaved somewhat differently from the other two holoenzymes in that it was inactivated by pCMB (but not NEM or IAA) following a pseudo-first order process. The titration of the two apoenzymes with [14C]NEM and that of the holo-short-chain acyl-CoA dehydrogenase with [14C]pCMB indicated that all three acyl-CoA dehydrogenases contain a single essential cysteine residue/subunit. In the inactivation of holo-medium-chain and holo-long-chain acyl-CoA dehydrogenases with methylmercury halide, the same essential cysteine residue was modified without perturbing or releasing the enzyme-bound FAD. The inactivations of the three holoenzymes by appropriate organic sulfhydryl reagents were prevented by prior incubation with substrate. These experimental results indicate that the essential cysteine residue is located in the vicinity of the FAD- and substrate-binding sites within the active center of the enzymes. It appears, however, that this cysteine residue does not participate directly in FAD binding.
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PMID:An essential cysteine residue located in the vicinity of the FAD-binding site in short-chain, medium-chain, and long-chain acyl-CoA dehydrogenases from rat liver mitochondria. 396 65

1. The oxidation of NADH and NADPH catalysed by the soluble supernatant from the hepatopancreas of Octopus vulgaris is due to a single enzyme, which has been purified approximately 100-fold. The enzyme reacts rapidly with potassium ferricyanide, and more slowly with 2,6-dichlorophenol-indophenol. No activity is obtained with oxygen, cytochrome c, lipoic acid, vitamin K(1), vitamin K(3), ubiquinone-30, p-benzoquinone, 2-p-iodophenyl-3-p-nitrophenyl-5-phenyltetrazolium chloride or methylene blue. 2. GSH, cysteine and mercaptoethanol stimulate the enzymic activity up to fivefold. GSSG is without any apparent effect. When stimulated by GSH the enzyme becomes sensitive to dicoumarol, which produces an inhibition competitive with respect to the activator. 3. The purified enzyme contains an acid-removable flavine component, which has been identified as FMN by spectrofluorimetry and chromatography in three solvent systems. After acid ammonium sulphate treatment the enzymic activity is lost, but it can be almost fully restored by incubation with FMN. FAD produces only a partial reactivation.
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PMID:Purification and properties of a soluble reduced nicotinamide-adenine dinucleotide (phosphate) dehydrogenase from the hepatopancreas of Octopus vulgaris. 417 22

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 flavoprotein mercuric reductase catalyzes the two-electron reduction of mercuric ions to elemental mercury using NADPH as an electron donor. It has now been purified from Pseudomonas aeruginosa PAO9501 carrying the plasmid pVS1. In this plasmid system, where the mer operon is on the transposon Tn501, mercuric reductase comprises up to 6% of the soluble cellular protein upon induction with mercurials. The purification is a rapid (two-step), high yield (80%) procedure. Anaerobic titrations of mercuric reductase with dithionite revealed the formation of a charge transfer complex with an absorbance maximum around 540 nm. Striking spectroscopic similarities to lipoamide dehydrogenase and glutathione reductase were observed. These two enzymes, which catalyze the transfer of electrons between pyridine nucleotides and disulfides, are flavoproteins which contain an oxidation-reduction-active cysteine residue at the active site. The expectation that mercuric reductase contains a similar electron acceptor was confirmed when it was shown that mercuric reductase has the capacity to accept four electrons per FAD-containing subunit, and that two thiols become kinetically titrable by 5,5'-dithiobis-(2-nitrobenzoate) upon reduction with NADPH. These are characteristic features of the disulfide reductase class of flavoproteins. Further similarities with at least one of these enzymes, lipoamide dehydrogenase, include the E/EH2 midpoint potential (-269 mV), fluorescence properties, and extinction coefficients of E and EH2. Preliminary observations relevant to an understanding of the mechanism of mercuric reductase are discussed.
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PMID:Mercuric reductase. Purification and characterization of a transposon-encoded flavoprotein containing an oxidation-reduction-active disulfide. 627

The nucleotide sequence of a 3614 base-pair segment of DNA containing the sdhA gene, encoding the flavoprotein subunit of succinate dehydrogenase of Escherichia coli, and two genes sdhC and sdhD, encoding small hydrophobic subunits, has been determined. Together with the iron-sulphur protein gene (sdhB) these genes form an operon (sdhCDAB) situated between the citrate synthase gene (gltA) and the 2-oxoglutarate dehydrogenase complex genes (sucAB): gltA-sdhCDAB-sucAB. Transcription of the gltA and sdhCDAB gene appears to diverge from a single intergenic region that contains two pairs of potential promoter sequences and two putative CRP (cyclic AMP receptor protein)-binding sites. The sdhA structural gene comprises 1761 base-pairs (587 codons, excluding the initiation codon, AUG) and it encodes a polypeptide of Mr 64268 that is strikingly homologous with the flavoprotein subunit of fumarate reductase (frdA gene product). The FAD-binding region, including the histidine residue at the FAD-attachment site, has been identified by its homology with other flavoproteins and with the flavopeptide of the bovine heart mitochondrial succinate dehydrogenase. Potential active-site cysteine and histidine residues have also been indicated by the comparisons. The sdhC (384 base-pairs) and sdhD (342 base-pairs) structural genes encode two strongly hydrophobic proteins of Mr 14167 and 12792 respectively. These proteins resemble in size and composition, but not sequence, the membrane anchor proteins of fumarate reductase (the frdC and frdD gene products).
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PMID:Nucleotide sequence encoding the flavoprotein and hydrophobic subunits of the succinate dehydrogenase of Escherichia coli. 638 59

Detergent-solubilized NADPH-cytochrome P-450 reductase was purified from porcine hepatic microsomes and compared to the rabbit enzyme isolated under identical conditions. The porcine enzyme had an equivalent specific activity toward cytochrome c compared to the rabbit enzyme. When analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the porcine enzyme exhibited a major band at Mr = 80,000 and two additional bands at Mr = 20,000 and 60,000. The 20-kDa fragment was shown to be the COOH-terminal portion of the protein which contains a hydrophobic sequence of 28 residues homologous to the pyrophosphate-binding portion of the FAD-binding protein p-hydroxybenzoate hydroxylase. The 60-kDa fragment corresponded to the NH2-terminal portion of the protein since this peptide and the intact protein have blocked NH2 terminal. The trypsin-solubilized porcine enzyme has an NH2-terminal sequence which is homologous to the equivalent trypsin-solubilized enzymes from rat and rabbit (80% sequence homology). Eight cysteine-containing peptides were isolated from a tryptic digest of the S-carboxymethylated pig enzyme. Significant sequence homology was not found between these peptides and other flavoproteins, except for one peptide (Glu-Val-Gly-Glu-Thr-Leu-Leu-Tyr-Tyr-Gly-Cys-Arg) which exhibited partial homology with the known NADPH-binding site of glutathione reductase. When the NADPH-protected enzyme was first S-alkylated with unlabeled iodoacetate, NADPH depleted, and further alkylated with 14C-labeled iodoacetate, the above radiolabeled peptide was isolated from a tryptic digest. The equivalent peptide was also isolated by a similar procedure from rabbit liver cytochrome P-450 reductase.
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PMID:Structural analysis of NADPH-cytochrome P-450 reductase from porcine hepatic microsomes. Sequences of proteolytic fragments, cysteine-containing peptides, and a NADPH-protected cysteine peptide. 643 80

3-Oxo-3-phenylpropyne and 3-oxo-3-phenylpropene were synthesized as active-site-directed irreversible inhibitors of the bitter almond hydroxynitrile lyase (EC 4.1.2.10), an FAD-protein. The substrate and competitors (e.g. benzoate) decrease the rate of the inhibitor-mediated deactivation of the enzyme. By excess addition of either one of the two inhibitors, the deactivation process is shown to be pseudo-first order. The reaction with equimolar amounts of 3-oxo-3-phenylpropyne with the enzyme is accompanied by a shift in the ultraviolet spectrum of the inhibitor, allowing direct measurement of the enzyme-inactivation process. The spectral change has second-order kinetics. Incubation with 3-oxo-3-[p-3H]phenylpropyne or 3-oxo-3-[1-14C]phenylpropene shows a one-to-one stoichiometry for the inhibitory-enzyme reaction. Dissociation of the 3-oxo-3[p-3H]phenylpropyne-inactivated holoenzyme with acid ammonium sulfate yields a labeled apoenzyme; the inhibitor does not react with free or enzyme-bound FAD. After boranate reduction and exhaustive hydrolysis of the 3-oxo-3-[1-14C]phenylpropene-inactivated enzyme, a labeled cysteine derivative was isolated which was identified by chromatographic and mass spectroscopic comparison with synthetic references as L-2-amino-4-thia-DL-7-hydroxy-7-phenylhepatanoic acid, the reduced, linear addition product of the inhibitor to a cysteine-SH group.
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PMID:Chemical modification of hydroxynitrile lyase by selective reaction of an essential cysteine-SH group with alpha, beta-unsaturated propiophenones as pseudo-substrates. 669 88

Sulphydryl oxidase is known to catalyse the synthesis de novo of disulphide bonds in a variety of thiol-containing compounds. Reduced glutathione is the best thiol substrate; however, D- and L-cysteine, cysteamine and N-acetyl-L-cysteine, as well as cysteine-containing peptides and proteins, are also effectively oxidized. In contrast, oxidation of the thiol groups of mercaptoethanol, mercaptopyridine, dithiothreitol, dithioerythritol, mercaptoacetate, mercaptopropionate or lipoic acid is not detectably catalysed. In bovine milk, sulphydryl oxidase is closely associated with another glutathione-metabolizing enzyme, gamma-glutamyltransferase. Covalent chromatography of crude preparations on cysteinylsuccinamidopropyl-glass resolves the oxidase from the transferase, thus permitting the kinetic characterization of glutathione oxidation. Initial-rate data imply a Ter Bi substituted-enzyme mechanism, and the observed substrate inhibition by thiols suggest that O2 binds first. Independent, non-kinetic, data, namely the immobilization of sulphydryl oxidase on cysteinyl-matrices, support formation of a mixed-disulphide intermediate between the thiol and enzyme, as predicted by the proposed mechanism. The enzyme-catalysed reaction appears not to be mediated via a superoxide intermediate, since O2 consumption is not affected by the presence of Nitro Blue Tetrazolium. FAD, NAD+, NADP+ and Nitro Blue Tetrazolium are all inactive as electron acceptors for sulphydryl oxidase catalysis.
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PMID:Kinetic mechanism and specificity of bovine milk sulphydryl oxidase. 674 73

Fumarate reductase, purified from the cytoplasmic membrane of Escherichia coli, has been cross-linked with the bifunctional reagent dimethylsuberimidate and shown to exist as an alpha beta dimer of polypeptides of molecular weights 69,000 and 25,000 in a 1:1 molar ratio. The protein has an s20,w of 7.67S and a D20,w of 6.5 X 10(-7) cm2/s. The purified enzyme contained 4-5 mol of nonheme iron and 4-5 mol of acid labile sulfur while the visible absorption spectrum showed a broad peak between 400 and 470 nm owing to the presence of an Fe-S centre and 8 alpha[N-3]histidyl FAD. Fumarate reductase activity was readily inhibited by the sulfhydryl reagents 5,5'-dithiobis-(2-nitrobenzoic acid), p-chloromercuribenzoate, and iodoacetamide. Using 5,5'-dithiobis-(2-nitrobenzoic acid) sulfhydryl group modification was followed as a function of enzyme activity. A single cysteine residue was shown to be required for activity and this essential sulfhydryl group was located in the 69,000 dalton subunit. The amino acid composition of E. coli fumarate reductase was similar to the succinate dehydrogenases from beef heart mitochondrion and Rhodospirillum rubrum.
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PMID:Molecular properties of fumarate reductase isolated from the cytoplasmic membrane of Escherichia coli. 675 4


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