Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P47989 (xanthine oxidase)
 8,633 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A selenium-containing catalytic antibody (Se-4A4), prepared by converting reactive serine residues of a monoclonal antibody (4A4) raised against a GSH derivative into selenocysteines, acts as a mimic of cytosolic glutathione peroxidase (cGPX). To clarify the mechanism of action of this catalytic antibody, detailed studies on kinetic behaviour and biological activity were carried out. A rate of acceleration (kcat/Km/kuncat) 10(7)-fold that of the uncatalytic reaction is observed. Under similar conditions, the turnover number (kcat) of Se-4A4 is 42% of that of the natural rabbit liver cGPX. The Se-4A4 reaction involves a Ping Pong mechanism, which is the same as that of the natural cGPX. The selenocysteine residue is located in the binding site of the antibody and is shown to be crucial for this activity. Of the thiol compounds tested, only GSH is able to serve as substrate for Se-4A4. It was demonstrated, using the free-radical-damage system (hypoxanthine/xanthine oxidase) of cardiac mitochondria, that Se-4A4 can protect mitochondria from free-radical damage at least 10(4)-fold more effectively than the natural cGPX.
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PMID:Biochemical characterization of selenium-containing catalytic antibody as a cytosolic glutathione peroxidase mimic. 957 75

It has been shown that erythrocyte membrane proteins become susceptible to degradation by membrane-bound serine protease activity after oxidative modification of the membranes (M. Beppu, M. Inoue, T. Ishikawa, K. Kikugawa, Biochim. Biophys. Acta 1196 (1994) 81-87). The aim of the present study was to clarify the presence of the serine protease in oxidized erythrocyte membranes and to characterize the selectivity of the enzyme to oxidized proteins. Human erythrocytes were oxidized in vitro with xanthine/xanthine oxidase/Fe(III) and oxidized membranes isolated. Proteolytic activity of the membranes toward spectrin obtained from oxidized membranes and bovine serum albumin oxidized with H2O2/horseradish peroxidase was increased by membrane oxidation, and the degradability of the substrates was increased by substrate oxidation. The proteolytic activity was inhibited by the serine protease inhibitor diisopropyl fluorophosphate (DFP). The 72 kDa and 80 kDa proteins in the membranes were labeled by [3H]DFP when detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing conditions and subsequent fluorography. The 72 kDa protein was found to be a serine enzyme, acetylcholine esterase. The 80 kDa protein appeared to be responsible for the degradation of oxidatively damaged proteins. The 80 kDa protein was loosely bound to membranes and readily solubilized into a 0.1% NP-40 detergent solution. The presence of the same 80 kDa protease in intact erythrocyte cytosol was suggested. The increased serine protease activity in oxidized membranes can result from the increased adherence of the cytosolic 80 kDa serine protease to the membranes due to oxidation.
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PMID:Characterization of membrane-bound serine protease related to degradation of oxidatively damaged erythrocyte membrane proteins. 981 51

Biodegradation of poly(urethane)s (PU)s using single enzymes in vitro was assessed by measuring radiolabel release from model poly(ester-urea-urethane) (PESU) and poly(ether-urea-urethane) (PETU) materials synthesized with 14C-labelled monomers. Cholesterol esterase (CE), an enzyme found in monocyte-derived macrophages (MDM), has been reported to cause a significant level of radiolabel release from both of these PUs. Previous work has shown that CE activity could be inhibited by the serine protease/esterase inhibitor, phenylmethylsulfonyl fluoride. Since many serine proteases are present in circulating blood and can be released by cells other than MDM, this study investigated the ability of serine proteases relative to that of CE to cause the degradation of PUs. In addition, the possible role of several oxidative enzymes in the breakdown of PUs was investigated. Proteinase K, chymotrypsin and thrombin, when incubated with PESU, coated on glass slips, caused significant radiolabel release, with proteinase K giving the highest values. However, the highest radiolabel release which proteinase K could elicit was ten times less than CE. Thrombin and then chymotrypsin were progressively worse in their biodegradative activity. Only CE, and not the serine proteases, could elicit a detectable radiolabel release from PETU. Although the release of reactive oxygen species and molecular oxygen occur around an implanted biomaterial, several oxidative systems (peroxidase, xanthine oxidase, catalase), known to produce one or more of these molecular species, were unable to induce radiolabel release from these PUs. The process of biodegradation as assessed by radiolabel release appears to be a specific hydrolytic process, while the role of oxidative enzymes remains less clear.
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PMID:The biodegradation of poly(urethane)s by the esterolytic activity of serine proteases and oxidative enzyme systems. 1042 27

The degradation of peroxisomal and nonperoxisomal proteins by endoproteases of purified peroxisomes from senescent pea (Pisum sativum L.) leaves has been investigated. In our experimental conditions, most peroxisomal proteins were endoproteolytically degraded. This cleavage was prevented, to some extent, by incubation with 2 mM phenylmethylsulfonylfluoride, an inhibitor of serine proteinases. The peroxisomal enzymes glycolate oxidase (EC 1.1.3.1), catalase (EC 1.11.1.6) and glucose-6-phosphate dehydrogenase (EC 1.1. 1.49) were susceptible to proteolytic degradation by peroxisomal endoproteases, whereas peroxisomal manganese superoxide dismutase (EC 1.15.1.1) was not. Ribulose-1,5-bisphosphate carboxylase/oxygenase (EC 4.1.1.39) from spinach and urease (EC 3.5. 1.5) from jack bean were strongly degraded in the presence of peroxisomal matrices. These results indicate that proteases from plant peroxisomes might play an important role in the turnover of peroxisomal proteins during senescence, as well as in the turnover of proteins located in other cell compartments during advanced stages of senescence. On the other hand, our data show that peroxisomal endoproteases could potentially carry out the partial proteolysis which results in the irreversible conversion of xanthine dehydrogenase into the superoxide-generating xanthine oxidase (EC 1. 1.3.22). This suggests a possible involvement of the peroxisomal endoproteases in a regulated modification of proteins.
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PMID:Proteolytic cleavage of plant proteins by peroxisomal endoproteases from senescent pea leaves 1050 97

Multiple enzymes may stimulate ROS production in VSMC and endothelial cells. These include NADH/NADPH oxidase, xanthine oxidase, lipoxygenases, cyclooxygenase, P-450 monooxygenases, and the enzymes of mitochondrial oxidative phosphorylation. In addition to generation of intracellular O2- by these enzymes, extracellular stimuli including lipophilic substrates, membrane permeant oxidants (e.g., H2O2), cytokines, and growth factors may modulate cellular redox state. Both intracellular and extracellular ROS act as second-messengers to activate tyrosine and serine-threonine kinases, such as the MAP kinase family. As discussed in the previous sections, regulation of the MAP kinases is one example of the complexity of ROS-dependent signal transduction. Although the complexity of ROS-mediated signal transduction is daunting, the diversity offers multiple therapeutic targets for pharmacologic intervention.
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PMID:Redox signals that regulate the vascular response to injury. 1060 87

The widely distributed xanthine oxidoreductase (XOR) system has been shown to be modulated upon exposure of animals to ionizing radiation through the conversion of xanthine dehydrogenase (XDH) into xanthine oxidase (XO). In the present work, radiomodification of the XOR system by phenylmethylsulfonyl fluoride (PMSF) and dithiothreitol (DTT) was examined using female Swiss albino mice which were irradiated with gamma rays at a dose rate 0.023 Gy s(-1). PMSF, a serine protease inhibitor, and DTT, the sulfhydryl reagent, were administered intraperitoneally prior to irradiation. The specific activities of XDH and XO as well as the XDH/XO ratio and the total activity (XDH+XO) were determined in the liver of the mice. The inhibition of XO activity, restoration of XDH activity, and increase in the XDH/XO ratio upon administration of PMSF were suggestive of irreversible conversion of XDH into XO mediated through serine proteases. The biochemical events required for the conversion were probably initiated during the early phase of irradiation, as the treatment with PMSF immediately after irradiation did not have a modulatory effect. Interestingly, DTT was not effective in modulating radiation-induced changes in the XOR system or oxidative damage in the liver of mice. The DTT treatment resulted in inhibition of the release of lactate dehydrogenase. However, the protection appears to be unrelated to the formation of TBARS. On the other hand, the presence of PMSF during irradiation inhibited radiation-induced oxidative damage and radiation-induced increases in the specific activity of lactate dehydrogenase. These findings suggest that a major effect of ionizing radiation is irreversible conversion of xanthine to xanthine oxidase.
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PMID:Radiomodfication of xanthine oxidoreductase system in the liver of mice by phenylmethylsulfonyl fluoride and dithiothreitol. 1085 70

Comparison of Hirosaki hairless rat (HHR) and Sprague-Dawley (SD) rat liver glutathione transferase (GST) subunits by HPLC revealed differences in subunit 3; a new peak was detected in HHR GSTs and this was tentatively named X. By chromatofocusing, the HHR GST form composed of peak X and SD rat GST 3-3 were eluted at pH 8.8 and 9.1 respectively. The former was more sensitive to the SH reagent N-ethylmaleimide (NEM) than the latter. GSSG treatment of peak X resulted in a shift of retention time (peak Y) by HPLC analysis. However, such conversion was not observed for the SD rat GST 3-3 following GSSG or dithiothreitol (DTT) treatment. Peak Y exhibited m/z values of 26091.9 and 26125.4 by matrix-assisted laser-desorption ionization-time-of-flight MS, higher than those of peak X by 304-307, equivalent to the molecular-mass value of GSH. On treatment with DTT, peak Y was converted into peak X, with release of a substance with HPLC-characteristics of GSH. This substance was confirmed to be GSH by liquid chromatography/MS. These results thus indicated peak Y to be a glutathionylated form of peak X. Quantification revealed the release of 4 nmol of GSH from 0.12 mg of the peak Y protein, corresponding to 4.8 nmol (M(r) 25000). The nucleotide sequence of HHR GST subunit 3 cDNA proved identical to that reported for pGTA/C44, possessing asparagine and cysteine as the 198th and 199th amino acid residues, respectively, corresponding to lysine and serine in subunit 3 of the SD rat. Thus peak X appeared to be the product of HHR GST subunit 3 cDNA. Treatment with N-(4-dimethylamino-3,5-dinitrophenyl)maleimide, a coloured analogue of NEM, followed by trypsin-treatment and sequencing of labelled peptides, identified the reactive cysteine residue of HHR GST subunit 3 to be located at position 199. Unlike SD rat GST 3-3, HHR GST 3-3 was not activated by treatment with xanthine and xanthine oxidase. These results suggest polymorphism of the rat GST subunit 3 gene with individual gene product variation in sensitivity to oxidative stress.
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PMID:Polymorphism of the glutathione transferase subunit 3 in Sprague-Dawley rats involves a reactive cysteine residue. 1094 54

Low-density lipoprotein (LDL) and its oxidized derivatives are hypothesized to impair vascular function by increasing superoxide anion (O.). To investigate mechanisms in situ, isolated carotid arteries were incubated with native LDL (nLDL) or minimally oxidized LDL (mmLDL). With the use of en face fluorescent confocal microscopy and hydroethidine, an oxidant-sensitive fluorescent probe, we found that nLDL increased O. in vascular endothelium greater than fourfold by an N(omega)-nitro-L-arginine methyl ester (L-NAME)-inhibitable mechanism. In contrast, mmLDL increased O. in vascular endothelium greater than eightfold by mechanisms that were partially inhibited by L-NAME and allopurinol and essentially ablated by diphenyleneiodium. These data indicate that both nLDL and mmLDL uncouple endothelial nitric oxide synthase (eNOS) activity and that mmLDL also activates xanthine oxidase and NADPH oxidoreductase to induce greater increases in O. generation than nLDL. Western analysis revealed that both lipoproteins inhibited A-23187-stimulated association of heat shock protein 90 (HSP90) with eNOS without inhibiting phosphorylation of eNOS at serine-1179 (phospho-eNOS), an immunological index of electron flow through the enzyme. As HSP90 mediates the balance of.NO and O. generation by eNOS, these data provide new insight into the mechanisms by which oxidative stress, induced by nLDL and mmLDL, uncouple eNOS activity to increase endothelial O. generation.
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PMID:Native LDL and minimally oxidized LDL differentially regulate superoxide anion in vascular endothelium in situ. 1212 24

Reactive oxygen species (ROS) and reactive nitrogen species (RNS) are signal-transducing molecules that regulate the activities of a variety of proteins. In the present investigation, we have compared the effects of superoxide (O2-), nitric oxide (NO), and hydrogen peroxide (H2O2) on the activities of three highly homologous serine/threonine phosphatases, protein phosphatase type 1 (PP1), protein phosphatase type 2A (PP2A), and calcineurin (protein phosphatase type 2B). Although superoxide, generated from xanthine/xanthine oxidase or paraquat, and NO, generated from (+/-)-(E)-4-ethyl-2-[(E)-hydroxyimino]-5-nitro-3-hexenamide or sodium nitroprusside, potently inhibited the phosphatase activity of calcineurin in neuroblastoma cell lysates, they had relatively little effect on the activities of PP1 or PP2A. In contrast, H2O2 inhibited the activities of all three phosphatases in lysates but was not a potent inhibitor for any of the enzymes. Calcineurin inactivated by O2-, NO, and H2O2 could be partially reactivated by the reducing agent ascorbate or by the thiol-specific reagent dithiothreitol (DTT). Maximal reactivation was achieved by the addition of both reagents, which suggests that ROS and RNS inhibit calcineurin by oxidizing both a catalytic metal(s) and a critical thiol(s). Reactivation of H2O2-treated PP1 also required the combination of both ascorbate and DTT, whereas PP2A required only DTT for reactivation. These results suggest that, despite their highly homologous structures, calcineurin is the only major Ser/Thr phosphatase that is a sensitive target for inhibition by superoxide and nitric oxide and that none of the phosphatases are sensitive to inhibition by hydrogen peroxide.
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PMID:Differential susceptibilities of serine/threonine phosphatases to oxidative and nitrosative stress. 1214 65

Bovine pulmonary artery smooth muscle possesses the tissue inhibitor of matrix metalloproteinase-2 (TIMP-2) as revealed by Western immunoblot study of its cytosol fraction with bovine polyclonal TIMP-2 antibody. This potent polypeptide inhibitor of matrix metalloproteinases (MMPs) was purified to homogeneity from cytosol fraction of bovine pulmonary artery smooth muscle. This inhibitor was purified by ammonium sulfate precipitation followed by gelatin sepharose and lentil lectin sepharose affinity chromatography and continuous elution electrophoresis by Prep Cell Model 491 (Bio-Rad, USA). SDS-PAGE revealed that the inhibitor has an apparent molecular mass of 21 kDa and was confirmed as TIMP-2 by (i) Western immunoblot assay using bovine polyclonal TIMP-2 antibody; and also by (ii) amino terminal amino acid sequence analysis of the purified inhibitor is found to be identical with TIMP-2 obtained from other sources. The purified 21 kDa inhibitor was found to be active against matrix metalloproteinase-2 (MMP-2, 72 kDa gelatinase) and matrix metalloproteinase-9 (MMP-9, 92 kDa gelatinase), the ambient MMPs in the pulmonary artery smooth muscle. The inhibitor was also found to be sensitive to the activated 72 kDa gelatinase-TIMP-2 complex and also active human interstitial collagenase. By contrast, it was found to be insensitive to the serine proteases: trypsin and plasmin. The inhibitor was heat and acid resistant and it had the sensitivity to trypsin degradation and reduction-alkylation. Treatment of the inhibitor with hydrogen peroxide, superoxide generating system (hypoxanthine plus xanthine oxidase) and peroxynitrite inactivated the inhibitor.
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PMID:Identification, purification and partial characterization of tissue inhibitor of matrix metalloproteinase-2 in bovine pulmonary artery smooth muscle. 1467 7


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