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)

Methods have been devised to examine the spectral properties and state of reduction of the pterin ring of molybdopterin (MPT) in milk xanthine oxidase and the Mo-containing domain of rat liver sulfite oxidase. The absorption spectrum of the native pterin was visualized by difference spectroscopy of each protein, denatured anaerobically in 6 M guanidine hydrochloride (GdnHCl), versus a sample containing the respective apoprotein and other necessary components. The state of reduction of MPT was also probed using 2,6-dichlorobenzenoneindophenol (DCIP) to measure reducing equivalents/MPT, after anaerobic denaturation of the protein in GdnHCl in the presence or absence of Hg2+. In the case of xanthine oxidase the data indicate that the terminal sulfide ligand of Mo causes the reduction of a native dihydro form of MPT to the tetrahydro level. This reduction does not occur if Hg2+ is added prior to denaturation of the protein. Based on its observed behavior, the native MPT in the Mo cofactor of xanthine oxidase is postulated to exist as a quinonoid dihydropterin. Quantitation of DCIP reduction by MPT of Mo fragment of sulfite oxidase showed a two-electron oxidation of MPT, even when the Mo fragment was denatured in the presence of Hg2+ to prevent internal reduction reactions due to sulfhydryls or sulfide. Difference spectra of DCIP-treated versus untreated Mo fragment showed that MPT had been fully oxidized. These data indicate that the native MPT in sulfite oxidase must be a dihydro isomer different from that in xanthine oxidase.
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PMID:The state of reduction of molybdopterin in xanthine oxidase and sulfite oxidase. 237 87

Inhibition of conversion from IMP to uric acid, which interferes with both spectrophotometric and radioisotopic assays of IMP dehydrogenase, by addition of allopurinol (0.1 mM), an inhibitor of xanthine oxidase, to the incubation system made it possible to determine the enzyme activity in crude liver extracts. With this improved assay method, the regulatory properties of the enzyme in crude extracts of liver and Yoshida sarcoma ascites cells were examined. In both tissues IMP dehydrogenase was found in the postmicrosomal supernatant. However, further centrifugation resulted in precipitation of the enzyme, the enzyme from Yoshida sarcoma ascites cells being precipitated more easily than that from rat liver. It was also found that IMP dehydrogenase activity increased during liver regeneration and that this increase was associated with the precipitate from the postmicrosomal fraction. These findings suggest that such a large sedimentable complex including IMP dehydrogenase might be formed in relation to cell growth. Most of the enzyme activity in rat liver and Yoshida sarcoma ascites cells was extracted in the supernatant obtained by centrifugation at 105,000 X g for 4 h after treatment of tissue homogenates with 1 M KCl, 0.75 M (NH4)2SO4, 2 M dimethylsulfoxide, 2 M KSCN, 25% glycerol, or 0.8 M guanidine-HCl. Treatment with 2% deoxycholate, 2% Triton X-100 or 2 M urea gave limited extraction. The enzyme was retained on a phenyl-Sepharose CL-6B or octyl-Sepharose CL-6B column and eluted with 0.8 M guanidine-HCl. These results suggested that the enzyme molecule has not only ionic but also hydrophobic domains, through which it interacts with other molecules of the enzyme itself and/or postmicrosomal cellular components.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:IMP dehydrogenase. I. Studies on regulatory properties of crude tissue extracts based on an improved assay method. 614 Feb 63

5-Ethynyluracil is a time-dependent and tight binding inhibitor of xanthine oxidase. The maximal value of the first-order rate constant for onset of inhibition is 0.01 s-1, and the concentration of 5-ethynyluracil which gives one-half of this value is 190 microM. Because the t1/2 for formation of active enzyme from inhibited enzyme is greater than 30 h in the absence of NADH, inhibition of xanthine oxidase by 5-ethynyluracil is functionally irreversible. One equivalent of 5-[2-14C]ethynyluracil/equivalent of active enzyme is required for complete inhibition. Allopurinol (100 microM), a potent inhibitor of xanthine oxidase, and cyanide (5 mM), an inactivator of the enzyme, do not abolish the binding of 5-[2-14C]ethynyluracil to the enzyme. Because radiolabel is released from 5-[2-14C]ethynyluracil-treated enzyme by treatment with 6 M guanidine HCl, a stable covalent bond is not formed between the inhibitor and the enzyme. However, the radiolabel released from inhibited enzyme is not 5-ethynyluracil. Moreover, NADH restores catalytic activity to the inhibited enzyme and displaces the radiolabel as 5-acetyluracil. Thermal denaturation of 5-ethynyluracil-inhibited xanthine oxidase results in the release of approximately equal amounts of 5-acetyluracil and a more hydrophilic product. Consequently, the 5-ethynyluracil-xanthine oxidase complex yields different degradation products of 5-ethynyluracil under different denaturation conditions. Seven uracil analogues with 5-substituents were tested as time-dependent inhibitors of xanthine oxidase. 5-Ethynyluracil is the only uracil analogue that potently inhibited xanthine oxidase. The reactivity of these uracil derivatives with sulfite was also determined. 5-Ethynyluracil is many fold more susceptible to nonenzymatic nucleophilic addition of sulfite than are the other analogues. Thus, the potency of these uracil analogues as inhibitors of xanthine oxidase is related to the nonenzymatic reactivity of the analogues with sulfite.
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PMID:Reaction of 5-ethynyluracil with rat liver xanthine oxidase. 796 25

Sodium-hydrogen exchange (NHE) represents an important process mediating myocardial ischemic and reperfusion injury, and NHE inhibitors have been shown to be effective cardioprotective agents against this form of injury. The precise mechanisms by which NHE inhibition protect the heart are not known and we therefore postulated that attenuation of oxidative stress could contribute to such protection. Accordingly, we examined whether the potent and specific NHE inhibitor 4-isopropyl-3-methylsulphonylbenzoyl-guanidine methanesulphonate (HOE 642, 5 microM) can protect isolated rat hearts against mechanical and biochemical impairment produced by either hydrogen peroxide (150 or 200 microM) or a free radical generating system consisting of purine (4.6 or 9.2 mM) and xanthine oxidase (20 or 40 U/L). HOE 642 significantly delayed and attenuated both the depression in left ventricular developed pressure (LVDP) as well as the elevation in left ventricular end-diastolic pressure (LVEDP) produced by both concentrations of hydrogen peroxide, although greater protection was generally seen against the lower hydrogen peroxide concentration, particularly with respect to LVEDP. Hydrogen peroxide, at both concentrations, significantly reduced high energy phosphate and glycogen contents and elevated lactate levels, all of which were significantly attenuated by HOE 642. In contrast, HOE 642 had no effect on functional impairment produced by either concentration of the free radical generating system. At its lower concentration, the combination of purine plus xanthine oxidase had no effect on energy metabolites, although a significant reduction in high energy phosphate stores was seen with the higher concentration. However, this was unaffected by HOE 642. The protective effect of HOE 642 was mimicked by another NHE inhibitor, methylisobutylamiloride (MIA, 5 microM). Our study therefore shows that NHE inhibition selectively protects against functional and metabolic impairment produced by hydrogen peroxide. Since hydrogen peroxide formation has been implicated in the development of ischemic and reperfusion injury, it is possible that the protective effect of NHE inhibition against this form of oxidative stress may explain in part the basis for the well-established salutary actions of NHE inhibitors in the ischemic and reperfused myocardium. Since HOE 642 failed to modify the response to free radical generators, it is unlikely that the protective effects of NHE inhibitors can be explained by a free radical scavenging mechanism.
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PMID:Effect of sodium-hydrogen exchange inhibition on functional and metabolic impairment produced by oxidative stress in the isolated rat heart. 919 59

A guanoxabenz [1-(2,6-dichlorobenzylideneamino)-3-hydroxyguanidine; an N-hydroxyguanidine] reducing enzymatic activity of rat spleen cytosol was investigated. By means of protein purification and N-terminal amino acid sequencing, the reducing activity was shown to reside in xanthine oxidase. The action of the enzyme on guanoxabenz resulted in the formation of guanabenz [1-(2,6-dichlorobenzylidene-amino)-3-guanidine]; the product formation could be monitored by HPLC and its identity was confirmed by NMR analysis. The reduction of guanoxabenz required xanthine or NADH as reducing substrates, while the process could be blocked by allopurinol, a selective inhibitor of xanthine oxidase. By using bovine milk xanthine oxidase, the guanoxabenz reducing activity of the enzyme was also verified. We conclude that guanoxabenz is a novel electron acceptor structure for xanthine oxidase.
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PMID:Identification of an N-hydroxyguanidine reducing activity of xanthine oxidase. 979 17

Guanoxabenz (1-(2,6-dichlorobenzylidene-amino)-3-hydroxyguanidine) and guanabenz (1-(2,6-dichlorobenzylidene-amino)-3-guanidine) are both known as centrally active antihypertensive drugs. We have previously shown that enzymatic activity in the rat spleen can induce N-reduction of guanoxabenz, leading to high affinity alpha 2-adrenoceptor binding, due to the formation of the alpha 2-adrenoceptor active drug, guanabenz. The spleen activity appears to reside in xanthine oxidase as it is activated by xanthine and blocked by allopurinol. We report that high affinity guanoxabenz binding is also induced in rat brain membranes after addition of NADH or NADPH cofactors. However, the brain process was clearly different from that of the spleen, as the formation of high affinity binding in the brain was not blocked by allopurinol. Moreover the NADH/NADPH activated mechanism of the brain membranes was not blocked by carbon monoxide and SKF525A, thus the activity appears not to reside in cytochrome P450 enzymes. Instead the activity was blocked by menadione and dicumarol. We conclude that the rat cerebral cortex contains an enzymatic activity that may activate guanoxabenz leading to formation of a metabolite showing high affinity for alpha 2-adrenoceptors. We also conclude that the rat brain activity is clearly distinct from that of the rat spleen.
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PMID:Characterization of guanoxabenz reducing activity in rat brain. 982 Aug 76

Oxidation by rat liver microsomes of 13 compounds involving a C=N(OH) function (including N-hydroxyguanidines, amidoximes, ketoximes, and aldoximes) was found to occur with the release of nitrogen oxides such as NO, NO2-, and NO3-. The greatest activities were observed with liver microsomes from dexamethasone-treated rats (up to 8 nmol of NO2- nmol of P450(-)1 min-1). A detailed study of the microsomal oxidation of some of these compounds was performed. Oxidation of N-(4-chlorophenyl)-N'-hydroxy-guanidine led to the formation of the corresponding urea and cyanamide in addition to NO, NO2-, and NO3-. Formation of all these products was dependent on NADPH, O2, and cytochromes P450. Oxidation of two arylamidoximes was found to occur with formation of the corresponding amides and nitriles in addition to nitrogen oxides. Oxidation of 4-(chlorophenyl)methyl ketone oxime gave the corresponding ketone and nitroalkane as well as NO, NO2-, and NO3-. These reactions were also dependent on cytochromes P450 and required NADPH and O2. Mechanistic experiments showed that microsomal oxidations of amidoximes to the corresponding nitriles and of ketoximes to the corresponding nitroalkanes are not inhibited by superoxide dismutase (SOD) and are performed by a cytochrome P450 active species, presumably the high-valent P450-iron-oxo complex. On the contrary, microsomal oxidation of N-hydroxyguanidines to the corresponding cyanamides was greatly inhibited by SOD and appeared to be mainly due to O2*- derived from the oxidase function of cytochromes P450. Similarly, microsomal oxidations of N-hydroxyguanidines and amidoximes to the corresponding ureas and amides were also found to be mainly performed by O2*-, as shown by the great inhibitory effect of SOD (70-100%) and the ability of the xanthine-xanthine oxidase system to give similar oxidation products. However, it is noteworthy that other species, such as the P450 Fe(II)-O2 complex, are also involved, to a minor extent, in the SOD-insensitive microsomal oxidative cleavages of compounds containing a C=N(OH) bond. Our results suggest a general mechanism for such oxidative cleavages of C=N(OH) bonds with formation of nitrogen oxides by cytochromes P450 and NO-synthases, with the involvement of O2*- and its Fe(III) complex [(FeIII-O2-) or (FeII-O2)] as main active species.
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PMID:Microsomal cytochrome P450 dependent oxidation of N-hydroxyguanidines, amidoximes, and ketoximes: mechanism of the oxidative cleavage of their C=N(OH) bond with formation of nitrogen oxides. 986 Aug 31

We here show that the novel N-hydroxyguanidine derivative PR5 (1-(3, 4-dimethoxy-2-chlorobenzylideneamino)-3-hydroxyguanidine) is acting as an alternative electron acceptor in xanthine oxidase catalyzed oxidation of xanthine. The reduction product is the corresponding guanidine derivative 1-(3, 4-dimethoxy-2-chlorobenzylideneamino)guanidine (PR9). The reaction occurs under both anaerobic and aerobic conditions. Moreover, EPR measurements show that the action of PR5 is associated with the inhibition of superoxide radical formation seen under aerobic conditions. PR5 also supports xanthine oxidase catalyzed anaerobic oxidation of NADH. Kinetic studies indicate that increasing xanthine concentration significantly increases the apparent K(m) of PR5, but it remains unaltered by changing NADH concentration. Moreover, the molybdenum center inhibitor allopurinol inhibits the PR5-sustained oxidation of xanthine and NADH equally well, whereas the flavin adenine dinucleotide site inhibitor diphenyliodonium (DPI) markedly inhibits only the PR5-sustained oxidation of NADH. We suggest that PR5 binds and becomes reduced at the molybdenum center of the xanthine oxidase. We also found that both PR5 and its reduction product PR9 can inhibit the oxygen-sustained xanthine oxidase reaction. The properties of PR5 suggest that it is a member of a novel class of compounds which we have termed xanthine oxidase electron acceptor-inhibitor drugs. The potential use of xanthine oxidase electron acceptor-inhibitors in the prevention of free radical mediated tissue damage in organ ischemia-reperfusion diseases is discussed.
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PMID:N-Hydroxyguanidine compound 1-(3,4-dimethoxy- 2-chlorobenzylideneamino)-3-hydroxyguanidine inhibits the xanthine oxidase mediated generation of superoxide radical. 1077 47

The major insecticide imidacloprid (IMI) is known to be metabolized by human cytochrome P450 3A4 with NADPH by imidazolidine hydroxylation and dehydrogenation to give 5-hydroxy-imidacloprid and the olefin, respectively, and by nitroimine reduction and cleavage to yield the nitrosoimine, guanidine, and urea derivatives. More extensive metabolism by human or rabbit liver microsomes with NADPH or rabbit liver cytosol without added cofactor reduces the IMI N-nitro group to an N-amino substituent, i.e., the corresponding hydrazone. A major metabolite on incubation of IMI in the human microsome-NADPH system is tentatively assigned by LC/MS as a 1,2,4-triazol-3-one derived from the hydrazone; the same product is obtained on reaction of the hydrazone with ethyl chloroformate. The hydrazone and proposed triazolone are considered here together (referred to as the hydrazone) for quantitation. Only a portion of the microsomal reduction and cleavage of the nitroimine substituent is attributable to a CYP450 enzyme. The cytosolic enzyme conversion to the hydrazone is inhibited by added cofactors (NAD > NADH > NADP > NADPH) and enhanced by an argon instead of an air atmosphere. The responsible cytosolic enzyme(s) does not appear to be DT-diaphorase (which is inhibited by several neonicotinoids), aldose reductase, aldehyde reductase, or xanthine oxidase. However, the cytosolic metabolism of IMI is inhibited by several aldo-keto-reductase inhibitors (i.e., alrestatin, EBPC, Ponalrestat, phenobarbital, and quercetin). Other neonicotinoids with nitroimine, nitrosoimine, and nitromethylene substituents are probably also metabolized by "neonicotinoid nitro reductase(s)" since they serve as competitive substrates for [(3)H]IMI metabolism.
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PMID:Neonicotinoid insecticides: reduction and cleavage of imidacloprid nitroimine substituent by liver microsomal and cytosolic enzymes. 1223 Apr 9

The guanidine compound ME10092 (1-(3,4-dimethoxy-2-chlorobenzylideneamino)-guanidine), which possesses a strong cardioprotective effect to ischemia-reperfusion, was assessed for different pharmacological actions that may underlie its cardioprotective effect. In the living rat ME10092 decreased the blood pressure and heart rate in a dose-dependent manner. We found ME10092 to bind to alpha 1- and alpha 2-adrenoreceptors with moderate affinity (Ki values 1-4 microM), and to block adrenaline-elicited contractile responses in isolated guinea pig aortas. Our results indicate that ME10092 possesses a certain anti-oxidant profile. Thus, in a competitive manner and with low affinity it inhibited the bovine milk xanthine oxidase enzyme, as well as NAD(P)H oxidase driven oxyradical formation in membrane fractions isolated from the rat brain. By using electron paramagnetic resonance we here show that, after its systemic administration, ME10092 modulates the nitric oxide (NO) content in several tissues of the rat in a time-dependent manner. However, in vitro ME10092 inhibited the activities of nitric oxide synthases nNOS and eNOS, but not that of iNOS. Our data give evidence that the cardioprotective effect of ME10092 could be mediated through pharmacological mechanisms that include some modulation of NO production, as well as possible inhibition of radical formation during ischemia-reperfusion.
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PMID:Investigations on the pharmacology of the cardioprotective guanidine ME10092. 1524 98


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