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Query: UNIPROT:P47989 (xanthine oxidase)
 8,633 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Studies by e.p.r. (electron-paramagnetic-resonance) spectroscopy and by stopped-flow spectrophotometry on turkey liver xanthine dehydrogenase revealed strong similarities to as well as important differences from the Veillonella alcalescens xanthine dehydrogenase and milk xanthine oxidase. The turkey enzyme is contaminated by up to three non-functional forms, giving molybdenum e.p.r. signals designated Resting I, Resting II and Slow. Slow and to a lesser extent Resting I signals are like those from the Veillonella enzyme, whereas Resting II is very like a resting signal described by K. V. Rajagopolan, P. Handler, G. Palmer & H. Beinert (1968) (J. Biol. Chem. 243, 3784-3796) for aldehyde oxidase. Another non-functional form that gives the Inhibited signal is produced on treatment of the enzyme with formaldehyde. Stopped-flow measurements at 450 nm show that, as for the milk enzyme, reduction by xanthine is rate-limiting in enzyme turnover. The active enzyme gives rise to Very Rapid and Rapid molybdenum(V) e.p.r. signals, as well as to an FADH signal. That these signals are almost indistinguishable from those of the milk enzyme, confirms the similarities between the active sites. There are two types of iron-sulphur centres that give signals like those in the milk enzyme, though with slightly different parameters. Quantitative reduction titration of the functional enzyme with xanthine revealed two important differences between the turkey and the milk enzymes. First, the turkey enzyme FADH/FADH2 system has a redox potential sufficiently low that xanthine is incapable of reducing the flavin completely. This finding presumably explains the very low oxidase activity. Secondly, whereas the Fe/S II chromophore in the milk enzyme has a relatively high redox potential, for the turkey enzyme the value of this potential is lower and similar to that of its Fe/S I chromophore.
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PMID:Studies by electron-paramagnetic-resonance spectroscopy and stopped-flow spectrophotometry on the mechanism of action of turkey liver xanthine dehydrogenase. 17 33

1. The mid-point reduction potentials of the various groups in xanthine oxidase from bovine milk were determined by potentiometric titration with dithionite in the presence of dye mediators, removing samples for quantification of the reduced species by e.p.r. (electron-paramagnetic-resonance) spectroscopy. The values obtained for the functional enzyme in pyrophosphate buffer, pH8.2, are: Fe/S centre I, -343 +/- 15mV; Fe/S II, -303 +/- 15mV; FAD/FADH-; -351 +/- 20mV; FADH/FADH2, -236 +/-mV; Mo(VI)/Mo(V) (Rapid), -355 +/- 20mV; Mo(V) (Rapid)/Mo(IV), -355 +/- 20mV. 2. Behaviour of the functional enzyme is essentially ideal in Tris but less so in pyrophosphate. In Tris, the potential for Mo(VI)/Mo(V) (Rapid) is lowered relative to that in pyrophosphate, but the potential for Fe/S II is raised. The influence of buffer on the potentials was investigated by partial-reduction experiments with six other buffers. 3. Conversion of the enzyme with cyanide into the non-functional form, which gives the Slow molybdenum signal, or alkylation of FAD, has little effect on the mid-point potentials of the other centres. The potentials associated with the Slow signal are: Mo(VI)/Mo(V) (Slow), -440 +/- 25mV; Mo(V) (Slow)/Mo(IV), -480 +/- 25 mV. This signal exhibits very sluggish equilibration with the mediator system. 4. The deviations from ideal behaviour are discussed in terms of possible binding of buffer ions or anti-co-operative interactions amongst the redox centres.
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PMID:Oxidation-reduction potentials of molybdenum, flavin and iron-sulphur centres in milk xanthine oxidase. 18 52

Redox potentials for the various centres in the enzyme xanthine dehydrogenase (EC 1.2.1.37) from turkey liver determined by potentiometric titration in the presence of mediator dyes, with low-temperature electron-paramagnetic-resonance spectroscopy. Values at 25 degrees C in pyrophosphate buffer, pH 8.2, are: Mo(VI)/Mo(V)(Rapid),-350 +/- 20mV; Mo(V) (Rapid)/Mo(IV), -362 +/- 20mV; Fe-S Iox./Fe-S Ired., -295 +/- 15mV; Fe-S IIox./Fe-S IIred., -292 +/- 15mV; FAD/FADH,-359+-20mV; FADH/FADH2, -366 +/- 20mV. This value of the FADH/FADH2 potential, which is 130mV lower than the corresponding one for milk xanthine oxidase [Cammack, Barber & Bray (1976) Biochem. J. 157, 469-478], accounts for many of the differences between the two enzymes. When allowance is made for some interference by desulpho enzyme, then differences in the enzymes' behaviour in titration with xanthine [Barber, Bray, Lowe & Coughlan (1976) Biochem. J. 153, 297-307] are accounted for by the potentials. Increases in the molybdenum potentials of the enzymes caused by the binding of uric acid are discussed. Though the potential of uric acid/xanthine (-440mV) is favourable for full reduction of the dehydrogenase, nevertheless, during turnover, for kinetic reasons, only FADH and very little FADH2 is produced from it. Since only FADH2 is expected to react with O2, lack of oxidase activity by the dehydrogenase is explained. Reactivity of the two enzymes with NAD+ as electron acceptor is discussed in relation to the potentials.
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PMID:Oxidation--reduction potentials of turkey liver xanthine dehydrogenase and the origins of oxidase and dehydrogenase behaviour in molybdenum-containing hydroxylases. 86 27

The mechanism of the enhancing effect of methyl viologen (MV) and flavin-adenine dinucleotide (FAD) on sulfoxide reduction which is mediated by a combination of aldehyde oxidase (AO) from guinea pig liver and one-electron reducing flavoenzymes, such as milk xanthine oxidase (XO), was examined. The activity of anaerobic reduction of diphenyl sulfoxide (DPSO) to diphenyl sulfide (DPS) was less than 1 nmol/min/mg protein of AO preparation in a system consisting of hypoxanthine, XO and AO. However, the sulfoxide reduction by this system was enhanced about 6- and 100-fold by the additions of FAD and MV, respectively. In the system containing MV or FAD, other one-electron reducing flavoenzymes such as nicotinamide adenine dinucleotide (reduced form) (NADH) dehydrogenase, lipoamide dehydrogenase and glutathione reductase with an appropriate electron donor, could replace XO. The ability of supplemented flavoenzymes to facilitate DPSO reduction correlated with their abilities to reduce MV and FAD. When AO was omitted from the combined system, no sulfoxide reduction was observed. Stoichiometric study revealed that MV semiquinone and FADH2 were oxidized at ratios of 2 and 1 mol, respectively, per mol of DPS formed. These results indicate that either MV or FAD serves as an electron carrier from the supplemented flavoenzymes to AO, a terminal reductase of sulfoxide.
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PMID:Sulfoxide reduction catalyzed by guinea pig liver aldehyde oxidase in combination with one-electron reducing flavoenzymes. 383 63

Product formation during the oxidation of xanthine oxidase has been examined directly by using cytochrome c peroxidase as a trapping agent for hydrogen peroxide and the reduction of cytochrome c as a measure of superoxide formation. When fully reduced enzyme is mixed with high concentrations of oxygen, 2 molecules of H2O2/flavin are produced rapidly, while 1 molecule of O2-/flavin is produced rapidly and another produced much more slowly. Time courses for superoxide formation and those for the absorbance changes due to enzyme oxidation were fitted successfully to the mechanism proposed earlier (Olson, J. S., Ballou, D. P., Palmer, G., and Massey, V. (1974) J. Biol. Chem. 249, 4363-4382). In this scheme, each oxidative step is initiated by the very rapid and reversible formation of an oxygen.FADH2 complex (the apparent KD = 2.2 X 10(-4) M at 20 degrees C, pH 8.3). In the cases of 6- and 4-electron-reduced enzyme, 2 electrons are transferred rapidly (ke = 60 s-1) to generate hydrogen peroxide and partially oxidized xanthine oxidase. In the case of the 2-electron-reduced enzyme, only 1 electron is transferred rapidly and superoxide is produced. The remaining electron remains in the iron-sulfur centers and is removed slowly by a second order process (ks = 1 X 10(4) M-1 s-1). When the pH is decreased from 9.9 to 6.2, both the apparent KD for oxygen binding and the rapid rate of electron transfer are decreased about 20-fold. This result is suggestive of uncompetitive inhibition and implies that proton binding to the enzyme-flavin active site affects primarily the rate of electron transfer, not the formation of the initial oxygen complex.
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PMID:The reaction of reduced xanthine oxidase with oxygen. Kinetics of peroxide and superoxide formation. 626 59

Rabbit liver aldehyde oxidase (AO), like milk xanthine oxidase (XO) and chicken liver xanthine dehydrogenase (XDH), possesses the following prosthetic groups: FAD, a functional Mo center, and two spectroscopically distinct iron-sulfur centers, one with gav less than 2.0 (termed Fe/S I) and the other with gav greater than 2.0 (termed Fe/S II) in the reduced enzyme. EPR spectra for the Mov species were found to be nearly identical in AO and XO for a number of enzyme complexes, and the midpoint reduction potentials for functional MoVI/MoV (-359 mV) and MoV/MoVI (-351 mV) were nearly the same in all three enzymes (50 mM phosphate, pH 7.8). A strong magnetic interaction between MoV and reduced Fe/S I, previously detected in XO and XDH, was also found in AO. No MoV-Fe/S II interaction could be detected in AO (nor in XO). In contrast, the order of reduction of Fe/S I and Fe/S II, as measured from their midpoint potentials, is reversed in AO (Em = -207 and -310 mV, respectively) as compared to XO (Em = -280 and -245 mV, respectively) in phosphate buffer at pH 7.8. The oxidized-reduced extinction coefficients at 450 and 550 nm for the two centers are also apparently reversed in AO and XO. Although magnetic interaction between FAD and one or both reduced Fe/S centers has been detected in both AO and XO, no magnetic interaction between the two reduced Fe/S centers themselves was found in AO (although such interaction has been seen in XO). The average FAD reduction potential is substantially more positive in AO (Em for FAD/FADH., -258 mV; FADH./FADH2, -212 mV at pH 7.8) than in XO or XDH. It can be concluded that although the properties and immediate environment of the functional Mo center are conserved in the three Mo hydroxylase enzymes, and all three enzymes possess the same set of prosthetic groups, the properties of the groups which transfer electrons from the Mo to the ultimate electron acceptor can vary substantially in AO, XO, and XDH.
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PMID:Properties of the prosthetic groups of rabbit liver aldehyde oxidase: a comparison of molybdenum hydroxylase enzymes. 628 79

Xanthine oxidase and xanthine dehydrogenase are enzymes involved in the metabolism of purines and pyrimidines in various organisms. Their relationship to one another has been the subject of considerable debate, primarily because of their proposed roles in ischemia/reperfusion damage in tissues. Differences in the kinetics and oxidation-reduction behavior of the two forms are accounted for by the presence in the dehydrogenase of a binding site for NAD+, as well as a substantially lower reduction potential for the flavin FADH./FADH2 couple of the dehydrogenase relative to the oxidase. This review presents recent advances of our understanding of the biochemistry and molecular biology of these systems, including a model for the overall morphology of xanthine oxidizing enzymes. The evidence that the two enzymes represent alternate forms of the same gene product, in some cases reversibly interconvertible between one another, is discussed.
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PMID:Flavoprotein structure and mechanism. 4. Xanthine oxidase and xanthine dehydrogenase. 764 15

Xanthine oxidase (XO) and xanthine dehydrogenase (XDH), two forms of the same enzyme isolated from cow's milk, have differing redox potentials of their chromophores. Both XDH and XO are capable of accepting 8 electrons per active site cluster of redox acceptors. By titrating XDH with redox indicator dyes of various potentials, the potentials have been determined for the flavin as well as for the 2Fe/2S centers of the enzyme at pH 7.5, 25 degrees C. The redox potential for the FAD/FADH. half-potential was found to be -270 +/- 5 mV and that for the FADH./FADH2 half potential, -410 +/- 5 mV. The first flavin half potential is close to the value which has been reported for XO (Porras, A. G., and Palmer, G. (1982) J. Biol. Chem. 257, 11617-11626). However, the second FAD half-potential is 180 mV lower in XDH than in XO, creating a 140-mV separation between the FAD potentials in XDH. This separation gives rise to a maximum development of the flavin semiquinone in XDH near 0.9 equivalent as confirmed by EPR quantitation of FADH. formed during reductive titrations. The potentials of both the 2Fe/2S centers in XDH were determined and found to be identical to the values which were found for the iron-sulfur centers in XO.
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PMID:Redox potentials of milk xanthine dehydrogenase. Room temperature measurement of the FAD and 2Fe/2S center potentials. 839 16