Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.17.1.4 (xanthine dehydrogenase)
 1,236 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The properties of the molybdenum iron-sulfur flavoprotein, aldehyde oxidase from rabbit livers, have been further investigated in comparison with bovine milk xanthine oxidase. In agreement with earlier work, the ultraviolet/visible spectra indicate that the flavin and iron-sulfur centres of the enzymes are quite similar to one another. The molybdenum centres have been compared by EPR spectroscopy of molybdenum(V) and regarding re-insertion of the sulfido ligand of molybdenum into the desulfo enzyme forms. The pH optimum for sulfide insertion is approximately 2 lower for aldehyde oxidase than for xanthine oxidase. A detailed comparison of molybdenum(V) EPR signals has been made for the signals known as Arsenite, Slow and Rapid. Computer simulation of spectra in 1H2O and 2H2O, at 9 and 35 GHz was used. Slow signals from the two enzymes are scarcely distinguishable from one another. Under the conditions used, aldehyde oxidase yielded only the Rapid type 2 signal, whereas xanthine oxidase gives both the Rapid type 1 and 2 signals. The nature of the structural difference between the Rapid type 1 and type 2 signal-giving species is discussed. It is concluded that the molybdenum centres of xanthine oxidase and aldehyde oxidase are indeed similar to one another and that such differences as exist between their molybdenum(V) EPR signals and re-sulfuration properties are related to differences only in the substrate-binding sites. N-terminal amino acid analyses have been performed on peptides obtained by trypsin cleavage of aldehyde oxidase. Comparison with a sequence previously deduced [Wright, R. M., Vaitaitis, G. M., Wilson, C. M., Repine, T. B., Terada, L. S. & Repine, J. E. (1993) Proc. Natl Acad. Sci. USA 90, 10690-10694] makes it clear that the latter is not, as was assumed, that of a xanthine dehydrogenase but of an aldehyde oxidase. In contrast to the situation with xanthine oxidase, attempts to convert non-proteolysed aldehyde oxidase to a dehydrogenase form by treatment with dithiothreitol were unsuccessful. The reason for this is considered in the light of sequence data in the literature. The location of the NAD(+)-binding site is discussed, and the sequence data are also discussed in relation to the molybdenum, iron-sulfur and substrate-binding sites.
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PMID:Properties of rabbit liver aldehyde oxidase and the relationship of the enzyme to xanthine oxidase and dehydrogenase. 755 19

The conversion of xanthine dehydrogenase to xanthine oxidase that produces oxygen radicals has been implicated in the ischemic injury to the myocardium and to the kidney. Xanthine dehydrogenase uses NAD as the electron acceptor to catalyze a reaction which does not produce any oxygen free radicals and may depress the conversion of xanthine dehydrogenase to xanthine oxidase. Nicotinamide is the preferred precursor for NAD. This study was conducted to examine the effect of an 18% casein diet supplemented with 0.5% nicotinamide on the activity of oxidoreductase and its two enzyme forms, xanthine dehydrogenase and xanthine oxidase, in kidney, heart and liver of female obese Zucker rats that spontaneously develop glomerulosclerosis, cardiomegaly and fatty liver. Lean litter mates were used as controls. Nicotinamide supplementation had no effect on the activities of these enzyme forms in the liver of either obese rats or lean rats. Obese rats fed the nicotinamide supplemented diet had higher activities of these enzyme forms in kidneys and hearts than unsupplemented diet fed obese rats, but this difference was not observed in lean rats. In unsupplemented rats, xanthine oxidase activity in the kidney was greater in lean rats than obese rats. Thus, the abnormalities observed in obese rats are unlikely attributable to the xanthine oxidase-mediated oxidant stress.
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PMID:Dietary nicotinamide supplementation increases xanthine oxidoreductase activity in the kidney and heart but not liver of obese Zucker rats. 761 99

Although mammalian xanthine oxidase exists originally as a dehydrogenase form in freshly prepared samples, it is converted to an oxidase form during purification, either irreversibly by proteolysis or reversibly by sulfhydryl oxidation of the protein molecule. However, avoiding proteolysis the mammalian enzyme can be purified as an interconvertible form and thus can be used to compare directly the properties of xanthine dehydrogenase and the oxidase derived from the same enzyme molecule. The cDNAs encoding the enzyme have been cloned from several sources, and structural information is becoming available. The most significant difference between the two forms is the protein conformation around FAD, which changes the redox potential of the flavin and the reactivity of FAD with the electron acceptors, NAD and molecular oxygen. The flavin semiquinone is thermodynamically stable in xanthine dehydrogenase, but is unstable in xanthine oxidase. Detailed analyses by stopped-flow techniques suggest that the flavin semiquinone reacts with oxygen to form superoxide anion while the fully reduced flavin reacts to form hydrogen peroxide. Although xanthine dehydrogenase can produce greater amounts of superoxide anion than xanthine oxidase during xanthine-oxygen turnover, it seems to be physiologically insignificant because NAD inhibits almost completely the formation of superoxide anion. Although the involvement of this enzyme in reperfusion injury has been proposed, this seems to be more complex than originally envisaged and still remains to be established.
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PMID:The conversion of xanthine dehydrogenase to xanthine oxidase and the role of the enzyme in reperfusion injury. 779 66

The amino acid sequence of chicken liver xanthine dehydrogenase (EC 1.1.1.204) was determined by cDNA cloning and partial amino acid sequencing of the purified enzyme. The enzyme consisted of 1358 amino acids with calculated molecular mass of 149,633 Da. In order to compare the structure of the chicken and rat enzymes, limited proteolysis was performed with the purified chicken liver xanthine dehydrogenase. When the enzyme was digested with subtilisin, it was not converted from the NAD-dependent dehydrogenase type to the O2-dependent oxidase type, in contrast with the mammalian enzyme. However, the enzyme was cleaved mainly into three fragments in a manner similar to that for the rat enzyme at pH 8.2 (20, 37, and 84 kDa) and retaining a full complement of redox centers. The cleavage sites were identified by determination of amino-terminal sequences of the produced fragments. It was concluded that the 20-kDa fragment was amino-terminal, the 84-kDa fragment carboxyl-terminal, and the 37-kDa fragment an intermediate portion in the enzyme protein. On the other hand, when the enzyme was digested with the same protease at pH 10.5, the sample contained only the 20- and 84-kDa portions and lacked the 37-kDa portion. The resultant sample possessed xanthine dichlorophenol indophenol reductase activity, indicating that the molybdenum center remained intact. The absorption spectrum showed the sample was very similar to deflavo-enzyme. From these results and sequence analyses, the domain structure of the enzyme is discussed.
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PMID:The structure of chicken liver xanthine dehydrogenase. cDNA cloning and the domain structure. 785 55

We have cloned and sequenced the hxA gene coding for the xanthine dehydrogenase (purine hydroxylase I) of Aspergillus nidulans. The gene codes for a polypeptide of 1363 amino acids. The sequencing of a nonsense mutation, hxA5, proves formally that the clones isolated correspond to the hxA gene. The gene sequence is interrupted by three introns. Similarity searches reveal two iron-sulfur centers and a NAD/FAD-binding domain and have enabled a consensus sequence to be determined for the molybdenum cofactor-binding domain. The A. nidulans sequence is a useful outclass for the other known sequences, which are all from metazoans. In particular, it gives added significance to the missense mutations sequenced in Drosophila melanogaster and leads to the conclusion that while one of the recently sequenced human genes codes for a xanthine dehydrogenase, the other one must code for a different molybdenum-containing hydroxylase, possibly an aldehyde oxidase. The transcription of the hxA gene is induced by the uric acid analogue 2-thiouric acid and repressed by ammonium. Induction necessitates the product of the uaY regulatory gene.
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PMID:Cloning and molecular characterization of hxA, the gene coding for the xanthine dehydrogenase (purine hydroxylase I) of Aspergillus nidulans. 787 88

The reductive half-reaction of milk xanthine dehydrogenase (XDH) with NADH and with xanthine has been studied at pH 7.5, 25 degree C. NADH reduces XDH to the two-electron reduced form at a rate of 18 s-1, independent of NADH concentration over the range studied. Further reduction by NADH to the four-electron state is inhibited by excess NADH. Subsequent binding of NADH to the four-electron reduced form of the enzyme causes the redistribution of one electron from the flavin to the molybdenum center. The four-electron reduced species reached through reduction by NADH is the same as the species obtained upon reaction of NAD with fully reduced XDH. In contrast, xanthine rapidly reduces XDH to the four-electron level; further reduction is comparatively slow and is inhibited by excess xanthine. Studies using substoichiometric xanthine show that the reaction of XDH with 1 equivalent of xanthine involves rapid substrate binding and rapid reduction of the molybdenum center of the enzyme. Before the release of urate from the molybdenum active site, an electron is transferred at 15 s-1 from the reduced molybdenum center to one of the iron-sulfur centers of XDH. Urate is then released at a rate of 13 s-1, followed by a rapid electron redistribution within the protein. The reductive half-reaction of XDH with xanthine is rate-limiting in xanthine/NAD turnover, which appears to occur between the two- and four-electron reduced enzyme species. The reduction of XDH by substoichiometric amounts of the fluorescent substrate xanthopterin was also studied. This reaction, monitored by changes in both absorbance and fluorescence, was found to involve the formation of two molybdenum complexes (an Eox.S complex and an Ered.P complex) followed by the release of the product, leucopterin.
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PMID:Studies of the reductive half-reaction of milk xanthine dehydrogenase. 803 47

These studies examined the kinetic and mechanistic parameters of mitomycin C (MMC) bioreduction by xanthine dehydrogenase (XDH), an enzyme recently shown to be capable of MMC activation. The bioreduction of MMC by XDH leads to the formation of 2,7-diaminomitosene (2,7-DM) under both aerobic and hypoxic conditions, with greater 2,7-DM formation observed under hypoxic conditions. The XDH-induced formation of 2,7-DM is pH dependent with increasing formation as the pH is varied from 7.4 to 6.0. In this study, the kinetics of MMC bioreduction by XDH was assessed under aerobic and hypoxic conditions and at pH 7.4 and 6.0. MMC interaction with XDH was also assessed by monitoring the ability of MMC to inhibit XDH-mediated uric acid and NADH formation. The ability of xanthine to serve as reducing equivalents for MMC reduction was also measured. Aerobically but not hypoxically, MMC reduction by XDH followed Michaelis-Menten kinetics. Kinetic constants calculated under aerobic conditions suggested that the pH-dependent increase (pH 6.0 > pH 7.4) in MMC activation by XDH is due to an approximately 2-fold decrease in the Km and a 2-fold increase in the Vmax at pH 6.0. Stimulation of uric acid formation and decreases in NADH formation by XDH in the presence of MMC suggest that MMC interaction with XDH may occur at the NAD(+)-binding region of the enzyme. The ability of xanthine to serve as reducing equivalents for MMC conversion to 2,7-DM also supports the hypothesis that MMC reduction is occurring at the NAD+ site.
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PMID:Kinetics and mechanism of mitomycin C bioactivation by xanthine dehydrogenase under aerobic and hypoxic conditions. 822 87

The reduction of milk xanthine dehydrogenase by salicylate anion radical (SL-), nicotinamide adenine dinucleotide radical (NAD.), and 1-methylnicotinamide (NMA) radicals was investigated by the use of pulse radiolysis. Reduction of the dehydrogenase with SL- proceeded via two phases. From the kinetic difference spectra obtained, the faster and slower phases of reduction represent that of one of the iron-sulfur centers and of FAD, respectively. The rate constant of the faster phase increased with the concentration of the enzyme, suggesting that the reduction follows a bimolecular reaction of SL- with the iron-sulfur center. In contrast, the rate constant of the slower phase (510 s-1) was independent of the concentration of the enzyme at pH 7.5. In order to elucidate the contribution of the molybdenum site in the reaction, a similar reaction was performed with enzyme modified with oxipurinol. In the modified enzyme, the slower phase was lost, whereas the faster phase was not affected. These results suggest that the slower phase is due to intramolecular electron transfer from the molybdenum center to FAD. On the other hand, NAD. reacted predominantly with FAD of the dehydrogenase to form the neutral semiquinone of FAD with a second order rate constant of 1.4 x 10(7) M-1 s-1 at pH 7.5, whereas a similar reaction in the oxidase, which was converted from xanthine dehydrogenase by proteolytical cleavage, was not observed. This suggests that NAD. transfers an electron via the binding site for NAD+ on the dehydrogenase. In contrast, NMA radical reduced only an iron-sulfur center of the dehydrogenase with a second order rate constant of 6.5 x 10(7) M-1 s-1 at pH 7.5.
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PMID:Electron transfer process in milk xanthine dehydrogenase as studied by pulse radiolysis. 822 23

We isolated cDNAs encoding xanthine dehydrogenase (XD; xanthine:NAD+ oxidoreductase, EC 1.1.1.204) from a human liver cDNA library. The complete nucleotide sequence of human XD was determined; the deduced amino acid sequence encoded a protein of 1336 amino acid residues of M(r) 147,782. Human XD possessed many of the signature sequences typical of XDs from flies and rodents, including an unusual cysteine distribution, a potential 2Fe/2S binding site, and a putative molybdopterin cofactor binding domain. Analysis of potential NAD binding sites suggested a simple hypothesis for the conversion of human XD into the oxygen metabolite forming xanthine oxidase (XO; xanthine:oxygen oxidoreductase, EC 1.1.3.22). Using a human XD complementary RNA hybridization probe, we found a 5100-base RNA in human liver by RNA blot-hybridization analysis. This RNA exhibited tissue-specific distribution that may be pertinent to XD- and XO-mediated oxygen radical injury in ischemia/reperfusion and inflammation. A second 4500-base RNA was detected in some tissues and may arise through differential transcription termination.
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PMID:cDNA cloning, characterization, and tissue-specific expression of human xanthine dehydrogenase/xanthine oxidase. 824 61

Mitomycin C (MMC), an alkylating anti-tumor agent, was activated by non-enzymatic and enzymatic mechanisms leading to DNA binding and adduct formation. However, it was enzymatically, not non-enzymatically, activated MMC which induced inter-strand DNA cross-linking, a major determinant of cell death. The enzymatic activation of MMC was catalyzed by microsomal NADPH:cytochrome P450 reductase (P450 reductase) and cytosolic enzyme activities. Human P450 reductase, transiently expressed from its cDNA in the COSI cells, metabolically activated MMC to generate 9 specific MMC-DNA adducts and induced inter-strand DNA cross-linking. Co-chromatography of the MMC-DNA adducts generated by P450 reductase and sodium borohydride in separate experiments indicated that MMC was metabolized by P450 reductase to produce 2,7-diaminomitosenes that exhibited binding to deoxyguanosine. Several experiments indicated that cytosolic enzymes which catalyzed reductive activation of MMC and DNA cross-linking included NAD(P)H:quinone oxidoreductaseI (NQOI or DT diaphorase) when present in extremely high concentrations and a unique cytosolic activity. The unique cytosolic activity was present in several mammalian cells and mouse colon and liver but absent in mouse kidney. The unique activity had properties of a diaphorase but was distinct from NQOI because of a lack of correlation between NQOI (2,6-dichlorophenolindophenol reduction) activity and the amount of MMC-reductive activation leading to DNA cross-linking. This activity was also distinct from xanthine oxidoreductase and NADH-cytochrome b5 reductase, 2 other enzymes that catalyze metabolic activation of MMC, because the unique activity was not inhibited by allopurinol (an inhibitor of xanthine oxidoreductase) and its activity was the same with NADH and NADPH (cytochrome b5 reductase is specific to NADH).
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PMID:Non-enzymatic and enzymatic activation of mitomycin C: identification of a unique cytosolic activity. 856 27


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