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)

Milk xanthine oxidase (XO) has been prepared in a dehydrogenase form (XDH) by purifying the enzyme in the presence of 2.5 mM dithiothreitol. Unlike XO, which reacts rapidly only with oxygen and not with NAD, the XDH form of the enzyme reacts rapidly with NAD. XDH has a turnover number for the NAD-dependent conversion of xanthine to urate of 380 mol/min/mol at pH 7.5, 25 degrees C, with a Km = < or = 1 microM for xanthine and a Km = 7 microM for NAD, but has very little O2-dependent activity. There is evidence that the two forms of the enzyme have different flavin environments: XDH stabilizes the neutral form of the flavin semiquinone and XO does not. Further, XDH binds the artificial flavin 8-mercapto-FAD in its neutral form, shifting the pK of this flavin by 5 pH units, while XO binds 8-mercapto-FAD in its benzoquinoid anionic form. XDH can be converted back to the XO form by the addition of three to four equivalents of the disulfide-forming reagent 4,4'-dithiodipyridine, suggesting that, in the XDH form of the enzyme, disulfide bonds are broken; this may cause a conformational change which creates a binding site for NAD and changes the protein structure near the flavin.
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PMID:Purification and properties of milk xanthine dehydrogenase. 132 33

Clinical evidence has suggested that mitomycin C (MMC) potentiates doxorubicin (DOX) induced cardiotoxicity. In this study a mouse model was used to examine the effect of DOX on the ability of cardiac tissue to bioactivate MMC to generate oxygen radicals. Cardiac damage was assessed by measuring serum CPK-MB isoenzyme levels and thiobarbituric acid reactive substances (TBARS) in the cardiac tissue. The exposure of animals to DOX or DOX and MMC over a three week period led to an increase in serum CPK-MB isoenzyme levels as well as TBARS. Treatment with DOX led to an increase in MMC-dependent, NADH-dependent, cyanide insensitive oxygen consumption, compared to control animals, thereby suggesting increased MMC-dependent oxygen radical generation. Levels of xanthine oxidase (XO; EC 1.1.3.22) and NADPH:cytochrome C reductase, two enzymes known to bioactivate MMC with subsequent oxygen radical generation, were measured in cardiac tissue with a 4.5 x increase in XO activity seen in DOX treated animals vs controls and no change in NADPH:cytochrome C reductase activity. Cardiac levels of xanthine dehydrogenase (XDH; EC 1.1.1.204) activity in DOX treated animals decreased while the XO/XDH ratio increased, suggesting a conversion of XDH to XO following DOX treatment.
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PMID:Role of xanthine oxidase in the potentiation of doxorubicin-induced cardiotoxicity by mitomycin C. 191 Oct 46

The present study tested the hypothesis that calpain is responsible for the limited proteolytic conversion of xanthine dehydrogenase (XD) to xanthine oxidase (XO). We compared the effects of various proteases on the activity and molecular weight of a purified preparation of xanthine dehydrogenase from rat liver. In agreement with previous reports, trypsin treatment produced a complete conversion of XD to XO accompanied by a limited proteolysis of XDH from an Mr of 140 kD to an Mr of 90 kD. Treatment with calpain I or calpain II did not produce a conversion from XD to XO nor did it result in partial proteolysis of the enzyme. Similarly, trypsin treatment partially degraded a reversibly oxidized form of xanthine dehydrogenase while calpain I or calpain II were ineffective. The possibility that an endogenous inhibitor prevented the proteolysis of XDH by calpain I or II was excluded by verifying that brain spectrin, a known calpain substrate, was degraded under the same incubation conditions. The results indicate that calpain is not likely to be responsible for the in vivo conversion of XD to XO under pathological conditions.
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PMID:Proteolytic conversion of xanthine dehydrogenase to xanthine oxidase: evidence against a role for calcium-activated protease (calpain). 255 23

An investigation was made into the possible involvement of the enzyme xanthine oxidase (XO) (EC 1.1.3.22), both reversible (XOrev) and irreversible (XOirr), in damage observed after short-term in vivo hepatic ischaemia/reperfusion (60 or 120 min I and 15 min R) in fasted rats with: (i) a physiological content of XO (25%); and (ii) higher XO percentage (45%). In the latter the hepatic XO physiological percentage was increased by diethylmaleate treatment (300 mg kg-1) that depleted the cytosolic glutathione (GSH) to 14% of the controls. It was shown that, in animals with physiological content of XO, 60 and 120 min of hepatic ischaemia followed by 15 min reperfusion results in decreased GSH levels, and significantly increased alanine aminotransferase (ALT) and aspartate aminotransferase (AST) serum levels, without any modification of either the percentages of XO (XOirr and XOrev) or the hepatic thiobarbituric acid reactive substances (TBARS). Sixty minutes of ischaemia/reperfusion in rats with the higher XO level and lower hepatic GSH content led to further conversion of XDH to XOrev, with no increase in XOirr. In addition, the ALT and AST serum levels in these animals rose to the same extent as in normal rats after 120 min ischaemia and 15 min reperfusion, this extent being observed to be associated with a moderate increase in thiobarbituric acid reactive substances (TBARS). However, the administration of allopurinol, at a dose of 50 mg kg-1, which almost completely inhibits XO activity, did not lead to any decrease in liver damage or TBARS.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:No documentable role for xanthine oxidase in the pathogenesis of hepatic in vivo ischaemia/reperfusion injury. 786 19

In the present study we have examined the potential role of xanthine oxidase (XO) in the intracellular oxidative stress induced by combinations of recombinant murine TNF alpha (rMuTNF alpha) and murine interferon-gamma (IFN gamma) in cultured mouse hepatocytes. IFN gamma alone and the combination of rMuTNF alpha and IFN gamma increased XO activity after a 4 hr exposure period. rMuTNF alpha alone increased XO activity only after 24 hr. At the later time point, the increased XO activity was accounted for by decreased XDH activity. However, the apparent conversion of XDH to XO cannot account for the early effects of rMuTNF alpha on hepatocyte function, particularly the onset of an oxidative stress (as indicated by efflux of GSSG from the hepatocytes). This effect is observed after two hours, and it is temporally the earliest sign of alteration of cellular function caused by rMuTNF alpha. Increased XO activity was not observed until 4 hr after treatment with rMuTNF alpha/IFN gamma. In addition, inhibition of XO activity with allopurinol did not ameliorate GSSG efflux from hepatocytes treated with the cytokines. However, the ATP depletion caused by the combination of rMuTNF alpha and IFN gamma and the cytotoxicity observed with the combined cytokines in cells pre-treated with BCNU (to inhibit glutathione reductase) was inhibited by allopurinol. These results show that the onset of oxidative stress in cultured mouse hepatocytes is not due to conversion of XDH to XO. However, events which follow the efflux of GSSG, such as ATP depletion and cytotoxicity in cells with impaired anti-oxidant defenses, may be partially due to increased XO activity, especially in cells treated with both rMuTNF alpha and IFN tau.
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PMID:The role of xanthine oxidase in oxidative damage caused by cytokines in cultured mouse hepatocytes. 796 49

The contribution of xanthine oxidoreductase (XDH + XO) to the extracellular release of hydrogen peroxide (H2O2) and intracellular H2O2 concentration in cultured bovine aortic endothelial cells (BAEC) was determined. Intracellular H2O2 concentration was measured by the aminotriazole-mediated inactivation of catalase, while extracellular H2O2 release was measured by the horse-radish peroxidase-mediated oxidation of p-hydroxyphenyl acetic acid to a fluorescent dimer. Supplementation of reaction systems with xanthine did not increase H2O2 production by cells. Inhibition of XO activity with allopurinol did not decrease either intracellular concentrations or the extracellular release of H2O2. Similarly, inactivation of XO by culture of cells with tungsten did not have any effect on intracellular levels of H2O2, while it increased extracellular release of H2O2 by 86 and 103% from cells cultured in Medium 199 (M199) and Dulbecco's modified Eagle's medium (DMEM), respectively. Cells cultured in DMEM had an average of 8 times greater XDH + XO specific activity, compared to M199 cultured cells, and had a threefold greater rate of release of H2O2 than M199-grown cells. However, DMEM-cultured cells did not have a greater rate of myxothiazole-resistant respiration, suggesting that this increase in H2O2 release comes from sources other than XO. These results show that cellular XO does not contribute significantly to basal H2O2 production in bovine endothelial cells. Analysis of XDH + XO activity of endothelial cells derived from vessels of various species showed a relatively low specific activity of this potential oxidant source in human-derived cells compared with cells cultured from other species such as rodents.
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PMID:Endogenous xanthine oxidase does not significantly contribute to vascular endothelial production of reactive oxygen species. 818 23

The release of xanthine oxidase/dehydrogenase XO/XDH (4.99 +/- 1.08 x 10(-12) moles/min/g wet wt in aerobic hearts) was not significantly increased following 30 min of ischemia, but almost doubled following 45 min of ischemia, indicating some endothelial cell damage. This release, however, was small compared to the release induced by digitonin perfusion (26 fold increase in an equivalent volume of effluent) and was nearly 1000 fold less than the enzyme activity measured in the tissue homogenate. There was a significant decrease in cardiac function (heart rate and systolic pressure) following 30 min of ischemia and no recovery of function following 45 min of ischemia and reperfusion. Endothelial cell damage determined by XO/XDH release is negligible during times of ischemia that cause severe damage to myocardial contractility. Coronary endothelial cells should not contribute significantly to circulating XO/XDH levels following ischemia.
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PMID:Xanthine oxidase/dehydrogenase release following ischemia in isolated rat hearts. 830 95

Xanthine dehydrogenase/xanthine oxidase (XDH/XO) is a major cytoplasmic source of superoxide radicals and hydrogen peroxide, and it is considered important in the pathogenesis of ischemia-reperfusion damage. Because little is known about the enzyme in human tissues, the aims of this study were to purify human XDH/XO and to produce Ab for detection of the protein in Western blots and for quantification by ELISA. We purified human milk XDH/XO, produced Ab for Western blotting and ELISA of the protein, and evaluated the molecular forms and activity-protein relationships in human tissues. The molecular size of the purified protein under nondenaturing conditions was approximately 300 kd. On SDS-PAGE, it was fragmented into four main bands of 143, 125, 87, and 59 kd. Ab recognized bands of similar size in Western blots of the purified preparation and human milk. In fresh liver homogenates treated with anti-proteases, the three largest bands were observed; in the intestine, only the two largest were observed. Serum, brain, heart, and skeletal muscle were negative, whereas some lung and kidney samples showed one faint band of 143 kd. Trypsin treatment of the enzyme converted the large molecular-weight bands into smaller bands, as did incubation of a liver homogenate without anti-proteases. XDH/XO protein concentrations (ng/mg total protein) were 146 +/- 70 in liver and 556 +/- 320 in intestine and less than 5 ng/ml in serum. The relationship of activity to protein (2.7-3.0 mumol/min/mg XDH/XO protein) was constant in liver and intestine during development. We conclude that 1) human XDH/XO has molecular size and subunit structure similar to other mammalian enzymes; 2) the polypeptide chain is unstable, also in the intact cell, despite retained activity; and 3) the amount of inactive XDH/XO in human liver and intestine is apparently small.
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PMID:Organ distribution and molecular forms of human xanthine dehydrogenase/xanthine oxidase protein. 856 97

During arousal from estivation in land snails, Otala lactea, active metabolic functions are restored within minutes and oxygen consumption increases dramatically. During the transition from the hypoxic conditions of estivation to normoxia it is possible that xanthine oxidase (XO) in hepatopancreas contributes to the observed lipid peroxidation. Using a fluorometric assay that is based on the oxidation of pterin, the activities and some properties of XO and XO+XDH (sum of XO and xanthine dehydrogenase activities) were measured in hepatopancreas extracts. Km values for pterin for XO and XO+XDH were 9 and 6 microM, respectively, and the Km of XDH for methylene blue was 5 microM. Both XO+XDH and XO activities were inhibited by allopurinol (I50 = 2 microM), pre-incubation at 40 degrees C, and by 5 min H2O2 pre-exposure. Inclusion of azide in the reaction promoted a rise of approximately 70-fold in the inactivation power of H2O2 due to inhibition of high endogenous catalase activity. The I50 for H2O2 of XO+XDH and XO activities in the presence of azide was 0.04 and 0.11 mM, respectively. Unlike the situation for mammalian XO, a previous reduction of O. lactea XO (by pterin) was not necessary to make the enzyme susceptible to H2O2 effects. Interestingly, methylene blue partially prevented both heat- and H2O2-induced inactivation of XO+XDH activity. These data indicate that the formation of an enzyme-methylene blue complex induces protection against heat and oxidative damage at the FAD-active site. Both XO and XO+XDH activites were significantly higher in snails after 35 days of estivation compared with active snails 24 h after arousal from dormancy. The ratio of XO/(XO+XDH) activities was also slightly increased in estivating O. lactea (from 0.07 to 0.09; P < 0.025). XO activity was 0.03 nmol.min-1.mg protein-1 in estivating snails. Compared with hepatopancreas catalase, XO activity is probably too low to contribute significantly to the net generation of oxyradicals, and hence to peroxidative damage. Rather, the low potential of XO to induce oxidative stress may constitute an adaptive advantage for O. lactea during arousal periods.
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PMID:Xanthine oxidase and xanthine dehydrogenase from an estivating land snail. 857 86

LPS and selected cytokines upregulate xanthine dehydrogenase/xanthine oxidase (XDH/XO) in cellular systems. However, the effect of these factors on in vivo XDH/XO expression, and their contribution to lung injury, are poorly understood. Rats were exposed to normoxia or hypoxia for 24 h after treatment with LPS (1 mg/kg) and IL-1beta (100 microg/kg) or sterile saline. Lungs were then harvested for measurement of XDH/XO enzymatic activity and gene expression, and pulmonary edema was assessed by measurement of the wet/dry lung weight ratio (W/D). Although treatment with LPS + IL-1beta or hypoxia independently produced a 2-fold elevation (p < 0. 05 versus exposure to normoxia and treatment with saline) in lung XDH/XO activity and mRNA, the combination of LPS + IL-1beta and hypoxia caused a 4- and 3.5-fold increase in these values, respectively. XDH/XO protein expression was increased 2-fold by hypoxia alone and 1.3-fold by treatment with LPS + IL-1beta alone or combination treatment. Compared with normoxic lungs, W/D was significantly increased by exposure to hypoxia, LPS + IL-1beta, or combination treatment. This increase was prevented by treatment of the animals with tungsten, which abrogated lung XDH/XO activity. In conclusion, LPS, IL-1beta, and hypoxia significantly upregulate lung XDH/XO expression in vivo. The present data support a role for this enzyme in the pathogenesis of acute lung injury.
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PMID:Upregulation of xanthine oxidase by lipopolysaccharide, interleukin-1, and hypoxia. Role in acute lung injury. 965 43


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