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: EC:1.17.3.2 (xanthine oxidase)
8,383 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In Klebsiella pneumoniae, NifL modulates the activity of the transcriptional activator NifA in response to combined nitrogen or external molecular oxygen. We recently showed that K. pneumoniae NifL is a flavoprotein which apparently senses oxygen through a redox-sensitive, conformational change. In order to study whether the nitrogen signal might be transmitted to NifA through a stable modification of NifL we characterized the redox properties of NifL synthesized in Escherichia coli in the presence of different nitrogen sources. FAD analyses showed that purified NifL carried FAD as cofactor independent of nitrogen and oxygen availability. The redox potential of NifL synthesized in the presence of ammonium was -277+/-5 mV at pH 8.0 and 25 degrees C, as determined by reduction with dithionite or with enzymatic reduction by xanthine oxidase in the presence of methyl viologen as redox mediator. When synthesized under nitrogen-limiting conditions, NifL showed a redox potential of -274+/-6 mV at pH 8.0 and 25 degrees C. Fully reduced NifL fractions, synthesized under either condition listed above, reoxidized rapidly in the presence of molecular oxygen. These results indicate that for NifL synthesized in E. coli, the redox potential of the NifL-bound FAD is not influenced by the nitrogen source. The two NifL fractions differed, however, in that a non-flavin specific absorbance at 420 nm was found only in NifL synthesized in the presence of ammonium.
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PMID:NifL of Klebsiella pneumoniae: redox characterization in relation to the nitrogen source. 1035 Jun 21

During inflammation and other pathological states, the lipid mediator platelet-activating factor (PAF) and reactive oxygen species (ROS) are both generated. We have been investigating the effect of exogenous PAF on ROS formation in the human keratinocyte cell line (HaCaT). ROS production, measured using luminol-enhanced chemiluminescence (CL), proved to be rapid, transient, PAF receptor-mediated, and totally dependent on an increase in intracellular Ca2+ ([Ca2+]i) and on the presence of extracellular Ca2+. Repeated administration of PAF resulted in refractoriness to the agonist in terms of both capacities to increase [Ca2+]i and generate ROS. The cells, however, continued to respond fully to other stimulants (bradykinin, epidermal growth factor, thapsigargin). The PAF-induced increases in [Ca2+]i (monitored using the fluorescent probe Fluo-3) were also rapid and transient and paralleled those of ROS generation. Relatively specific inhibitors of potential ROS-producing systems were administered in an attempt to characterize the ROS producing system(s). Inhibitors of xanthine oxidase, phospholipase A2, lipoxygenase, cyclooxygenase and NO synthase did not interfere with PAF evoked ROS. The flavoprotein inhibitor diphenyleneiodonium and the mitochondrial cytochrome oxidase inhibitor KCN, prevented generation of ROS, making NAD(P)H a candidate for the electron source of the ROS and the mitochondria a potential major site of formation.
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PMID:Calcium-dependent PAF-stimulated generation of reactive oxygen species in a human keratinocyte cell line. 1036 77

Cell-free extract prepared from a mixed culture consisting of strains belonging to the genera Klebsiella and Rhodococcus grown in the presence of caffeine contains a novel enzyme, caffeine (1,3, 7-trimethylxanthine) oxidase which catalyzes the oxidation of caffeine at the C-8 position to produce 1,3,7-trimethyluric acid. The enzyme was purified to homogeneity by a combination of ion-exchange and hydrophobic column chromatographies. Both native and SDS/PAGE of the purified enzyme showed a single protein band and the subunit molecular mass of the protein was determined to be 85 kDa. Dichlorophenol indophenol and cytochrome c served as good electron acceptors but NAD and NADP did not. Caffeine served as the best substrate with an apparent K(m) of 11.4 microM. various analogues of theobromine were also effective substrates for caffeine oxidase. The activity was inhibited by o-phenanthroline, H(2)O(2), and methanol, but salicylate, thiol-group blocking reagents, and sodium arsenite, the known xanthine oxidase inhibitors, did not inhibit the reaction. The spectral characteristics of the purified enzyme suggest that it is a flavoprotein containing non-heme iron.
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PMID:Purification and partial characterization of caffeine oxidase--A novel enzyme from a mixed culture consortium. 1049 16

Although a burst of oxidants has been well described with reperfusion, less is known about the oxidants generated by the highly reduced redox state and low O(2) of ischemia. This study aimed to further identify the species and source of these oxidants. Cardiomyocytes were exposed to 1 h of simulated ischemia while oxidant generation was assessed by intracellular dihydroethidine (DHE) oxidation. Ischemia increased DHE oxidation significantly (0.7 +/- 0.1 to 2.3 +/- 0.3) after 1 h. Myxothiazol (mitochondrial site III inhibitor) attenuated oxidation to 1.3 +/- 0.1, as did the site I inhibitors rotenone (1.0 +/- 0.1), amytal (1.1 +/- 0.1), and the flavoprotein oxidase inhibitor diphenyleneiodonium (0.9 +/- 0.1). By contrast, the site IV inhibitor cyanide, as well as inhibitors of xanthine oxidase (allopurinol), nitric oxide synthase (nitro-L-arginine methyl ester), and NADPH oxidase (apocynin), had no effect. Finally, DHE oxidation increased with Cu- and Zn-containing superoxide dismutase (SOD) inhibition using diethyldithiocarbamate (2.7 +/- 0.1) and decreased with exogenous SOD (1.1 +/- 0.1). We conclude that significant superoxide generation occurs during ischemia before reperfusion from the ubisemiquinone site of the mitochondrial electron transport chain.
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PMID:Generation of superoxide in cardiomyocytes during ischemia before reperfusion. 1060 Aug 42

In this article, we report on the chromosome mapping and molecular cloning of the genetic locus encoding the mouse molybdo-iron/sulfur-flavoprotein aldehyde oxidase. The aldehyde oxidase locus maps to mouse chromosome 1 band C1-C2, as determined by fluorescence in situ hybridization experiments conducted on metaphase chromosomes. The gene is approximately 83 kb long and consists of 35 exons. The exon/intron boundaries are perfectly conserved relative to the corresponding human homolog and almost completely conserved relative to the mouse xanthine oxidoreductase gene. This further supports the concept that the aldehyde oxidase and xanthine oxidoreductase loci evolved from the same ancestral precursor by a gene duplication event. The position of a major transcription start site was defined by primer extension and RNase mapping analysis. The 5'-flanking region of the mouse aldehyde oxidase gene contains a functional and orientation-dependent promoter as well as several putative binding sites for known cell-specific and general transcription factors. Deletion analysis of the 5'-flanking region defines an approximately 470 bp DNA stretch which is necessary and sufficient for the transcription of the mouse aldehyde oxidase gene.
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PMID:The mouse aldehyde oxidase gene: molecular cloning, chromosomal mapping and functional characterization of the 5'-flanking region. 1067 24

Some sterically hindered N-substituted derivatives of daunorubicin are known to be poor substrates for NADH dehydrogenase, NADPH cytochrome P450 reductase and xanthine oxidase. In consequence, poor oxygen radical generation by these compounds is observed. In this study we examined a new family of sugar-N-substituted derivatives of daunorubicin bearing a bulky substituent introduced on the nitrogen atom through the amidine spacer. These compounds were found to be very active in radical formation catalyzed by all three studied enzymes. Thus, the introduction of a heterocyclic ring, even if it is bulky but flexible, on the nitrogen atom of daunosamine moiety through the one-atom spacer (amidine group), does not induce the steric hindrance effect on the interaction of daunorubicin derivatives with these flavoprotein enzymes.
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PMID:The ability of new formamidine sugar-modified derivatives of daunorubicin to stimulate free radical formation in three enzymatic systems: NADH dehydrogenase, NADPH cytochrome P450 reductase and xanthine oxidase. 1096 87

Glutaryl-CoA dehydrogenase catalyzes the oxidation of glutaryl-CoA to crotonyl-CoA and CO(2) in the mitochondrial degradation of lysine, hydroxylysine, and tryptophan. We have characterized the human enzyme that was expressed in Escherichia coli. Anaerobic reduction of the enzyme with sodium dithionite or substrate yields no detectable semiquinone; however, like other acyl-CoA dehydrogenases, the human enzyme stabilizes an anionic semiquinone upon reduction of the complex between the enzyme and 2,3-enoyl-CoA product. The flavin potential of the free enzyme determined by the xanthine-xanthine oxidase method is -0.132 V at pH 7.0, slightly more negative than that of related flavoprotein dehydrogenases. A single equivalent of substrate reduces 26% of the dehydrogenase flavin, suggesting that the redox equilibrium on the enzyme between substrate and product and oxidized and reduced flavin is not as favorable as that observed with other acyl-CoA dehydrogenases. This equilibrium is, however, similar to that observed in isovaleryl-CoA dehydrogenase. Comparison of steady-state kinetic constants of glutaryl-CoA dehydrogenase with glutaryl-CoA and the alternative substrates, pentanoyl-CoA and hexanoyl-CoA, suggests that the gamma-carboxyl group of glutaryl-CoA stabilizes the enzyme-substrate complex by at least 5.7 kJ/mol, perhaps by interaction with Arg94 or Ser98. Glu370 is positioned to function as the catalytic base, and previous studies indicate that the conjugate acid of Glu370 also protonates the transient crotonyl-CoA anion following decarboxylation [Gomes, B., Fendrich, G. , and Abeles, R. H. (1981) Biochemistry 20, 3154-3160]. Glu370Asp and Glu370Gln mutants of glutaryl-CoA dehydrogenase exhibit 7% and 0. 04% residual activity, respectively, with human electron-transfer flavoprotein; these mutations do not grossly affect the flavin redox potentials of the mutant enzymes. The reduced catalytic activities of these mutants can be attributed to reduced extent and rate of substrate deprotonation based on experiments with the nonoxidizable substrate analogue, 3-thiaglutaryl-CoA, and kinetic experiments. Determination of these fundamental properties of the human enzyme will serve as the basis for future studies of the decarboxylation reaction which is unique among the acyl-CoA dehydrogenases.
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PMID:Proton abstraction reaction, steady-state kinetics, and oxidation-reduction potential of human glutaryl-CoA dehydrogenase. 1098 95

Xanthine oxidase (EC 1.1.3.22) and xanthine dehydrogenase (EC 1.1.1. 204) are both members of the molybdenum hydroxylase flavoprotein family and represent different forms of the same gene product. The two enzyme forms and their reactions are often referred to as xanthine oxidoreductase (XOR) activity. Physiologically, XOR is known as the rate-limiting enzyme in purine catabolism but has also been shown to be able to metabolize a number of other physiological compounds. Recent studies have also demonstrated its ability to metabolize xenobiotics, including a number of anticancer compounds, to their active metabolites. During the past 10 years, evidence has mounted to support a role for XOR in the pathophysiology of inflammatory diseases and atherosclerosis as well as its previously determined role in ischemia-reperfusion injury. While significant progress has recently been made in our understanding of the physiological and biochemical nature of this enzyme system, considerable work still needs to be done. This paper will review some of the more recent work characterizing the interactions and the factors that influence the interactions of XOR with various physiological and xenobiotic compounds.
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PMID:Cellular distribution, metabolism and regulation of the xanthine oxidoreductase enzyme system. 1115 41

We report the cloning of the AOH1 and AOH2 genes, which encode two novel mammalian molybdo-flavoproteins. We have purified the AOH1 protein to homogeneity in its catalytically active form from mouse liver. Twenty tryptic peptides, identified or directly sequenced by mass spectrometry, confirm the primary structure of the polypeptide deduced from the AOH1 gene. The enzyme contains one molecule of FAD, one atom of molybdenum, and four atoms of iron per subunit and shows spectroscopic features similar to those of the prototypic molybdo-flavoprotein xanthine oxidoreductase. The AOH1 and AOH2 genes are 98 and 60 kilobases long, respectively, and consist of 35 coding exons. The AOH1 gene has the potential to transcribe an extra leader non-coding exon, which is located downstream of exon 26, and is transcribed in the opposite orientation relative to all the other exons. AOH1 and AOH2 map to chromosome 1 in close proximity to each other and to the aldehyde oxidase gene, forming a molybdo-flavoenzyme gene cluster. Conservation in the position of exon/intron junctions among the mouse AOH1, AOH2, aldehyde oxidase, and xanthine oxidoreductase loci indicates that these genes are derived from the duplication of an ancestral precursor.
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PMID:Purification of the aldehyde oxidase homolog 1 (AOH1) protein and cloning of the AOH1 and aldehyde oxidase homolog 2 (AOH2) genes. Identification of a novel molybdo-flavoprotein gene cluster on mouse chromosome 1. 1156 61

Xanthine oxidase (XO) is a highly versatile flavoprotein enzyme, ubiquitous among species (from bacteria to human) and within the various tissues of mammals. The enzyme catalyses the oxidative hydroxylation of purine substrates at the molybdenum centre (the reductive half-reaction) and subsequent reduction of O(2) at the flavin centre with generation of reactive oxygen species (ROS), either superoxide anion radical or hydrogen peroxide (the oxidative half-reaction). Many diseases, or at least symptoms of diseases, arise from a deficiency or excess of a specific metabolite in the body. For an example of an excess of a particular metabolite that produces a disease state is the excess of uric acid which can led to gout. Inhibition of XO decreases the uric acid levels, and results in an antihyperuricemic effect. Allopurinol, first synthesised as a potential anticancer agent, is nowadays a clinically useful xanthine oxidase inhibitor used in the treatment of gout. There is overwhelming acceptance that xanthine oxidase serum levels are significantly increased in various pathological states like hepatitis, inflammation, ischemia-reperfusion, carcinogenesis and aging and that ROS generated in the enzymatic process are involved in oxidative damage. Thus, it may be possible that the inhibition of this enzymatic pathway would be beneficial. In this review the State of the Art will be presented, which includes a summary of the progress made over the past years in the knowledge of the structure and mechanism of the enzyme, associated pathological states, and in the efforts made towards the development of new xanthine oxidase inhibitors.
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PMID:Progress towards the discovery of xanthine oxidase inhibitors. 1186 Mar 55


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