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)

An easily performed assay to identify the C3b and Fc receptors on human neutrophils was developed. Salmonella typhimurium were treated with fluorescein and then incubated in nonimmune fresh human serum, which led to C3b fixation via activation of the alternative pathway. Similarly, type II pneumococci were treated with fluorescein and opsonized with type-specific rabbit antiserum. Neutrophils bearing C3b and Fc receptors formed rosettes with the respective bacteria, which were easily readable because of their bright fluorescence. Incubation of neutrophils at 37 degrees C with C3-coated bacteria generated 54 +/ 4% C3b rosettes, whereas neutrophils incubated with immunoglobulin G-coated bacteria yielded 75 +/ 7% rosettes. Incubation at 4 degrees C inhibited the formation of C3b rosettes but not Fc rosettes. Heat inactivation of the fresh human serum at 56 degrees C for 30 min completely inhibited the formation of the C3b rosettes, and addition of heat-aggregated immunoglobulin G to the polymorphonuclear leukocyte blocked the ability of the polymorphonuclear leukocyte to bind immunoglobulin G-coated bacteria. Addition of 1.0 mM N-ethylmaleimide, 0.1 mg of trypsin per ml, 10 mM H2O2, O2- generated by xanthine-xanthine oxidase, and 8 times 10(-4) M hydrocortisone inhibited the C3b receptor, but did not inhibit the Fc receptor. In neutrophils, the selective effect of the various inhibitors suggests that the Fc and C3b receptors are distinct entities.
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PMID:Effects of surface-active agents on neutrophil receptors. 3 Jun 96

Two types of complexes are formed during the interaction of xanthine oxidase with p-chloromercurybenzoate (pCMB). The reversible inactive complex (presumably of absorption nature) is formed practically instantaneously and competitively with regard to the substrate (Ki=6,2 . 10(-8) M) in 0,05 M phosphate buffer (pH 7,8, 25 degrees) and does not involve the fast-reacting SH-groups of the enzyme. Reactivation of xanthine oxidase is observed during prolonged incubation of the inactive complex at 0 degrees; it is associated with the interaction between pCMB and the fact-reacting SH-groups. This interaction results in a dissociation of the inactive complex. The blocking of the slow-reacting SH-groups is accompanied by an irreversible loss of the xanthine oxidase activity. The enzyme modification by blocking of 10 fast-reacting SH-groups does not involve the Fe-S clusters, but results in local changes in the enzyme conformation. This is manifested in a 2-fold increase of Km and the rate constants of proteolysis of the modified xanthine oxidase as compared to the native enzyme. The rate constants of proteolysis by trypsin for the native and modified enzymes in 0,05 M phosphate buffer (pH 7,8; 37 degrees) are 3,7 . 10(-3) min-1 and 7,0 . 10(-3) min-1, respectively; those for chymotrypsin in the same buffer (30 degrees) are 1,5 . 10(-2) min-1 and 6,0 . 10(-2) min-1, respectively.
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PMID:[Mechanism of interaction between milk xanthine oxidase and p-chloromercuribenzoate. Properties of the purified enzyme]. 49 85

The heat stability at 56 C of xanthine oxidase (Xox) from mouse intestine was found to be affected by two factors: (1) Xox which had been partially digested by trypsin was less heat stable than Xox which was protected from digestion by the presence of soybean trypsin inhibitor and phenylmethylsulfonyl fluoride; (2) Xox was less heat stable in glass tubes than in polystyrene tubes and siliconized glass tubes resembled polystyrene tubes in this respect.
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PMID:Two factors affecting the heat stability of xanthine oxidase in extracts of mouse intestine. 120 60

Bovine milk xanthine oxidase (xanthine:oxygen oxidoreductase, EC 1.2.3.2) has been purified by a modified method without the use of proteases, and its structure has been analyzed by polyacrylamide gel electrophoresis. Native xanthine oxidase is found to consist of only two polypeptide chains A with molecular weights of 150 000 each. These chains have NH2-terminal methionine. Limited proteolysis with trypsin, chymotrypsin, or subtilisin at pH 8 did not affect molecular weight and activities of the enzyme while each of the A chains was cleaved under these conditions to three fragments C, E, and F with molecular weights of 92 00, 42 000 and 20 000, respectively. These fragments remained bound to each other and were relatively resistant to subsequent proteolysis. The isolation of xanthine oxidase in the presence of pancreatin as described by Hart et al. (1970, Biochem. J. 116, 851) gives partially digested enzyme composed mainly of chains C, E (Mr 35 000) and a small component (Mr approx. 15 0-0). The action of subtilisin on xanthine oxidase at pH 11 resulted in complete digestion of E chains, FAD separation, and total loss of xanthine:oxygen oxidoreductase activity while xanthine:indophenol oxidoreductase activity was relatively little affected. The residual enzyme has a molecular weight of about 200 000, is composed mainly of two C chains (and may probably contain F and/or proteolytic fragments of low molecular weight), contains molybdenum, and does not contain FAD.
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PMID:Subunit structure of bovine milk xanthine oxidase. Effect of limited cleavage by proteolytic enzymes on activity and structure. 126 10

Oxidative damage to bovine serum albumin (BSA) was induced by hydroxyl radical (HO.) generating systems of xanthine oxidase (XO) + EDTA-Fe3+ and ascorbate + EDTA-Fe3+. Formation of bityrosine and loss of tryptophan were observed in the ascorbate + EDTA-Fe3+ system and carbonyl formation was induced by both systems. Mannitol and ethanol very strongly inhibited the carbonyl and/or bityrosine formation, indicating that the oxidative damage to BSA was due to HO(.). The sulfhydryl (SH) groups of BSA were very sensitive to the XO + EDTA-Fe3+ but not to the ascorbate + EDTA-Fe3+ system. Catalase but not hydroxyl radical scavengers or superoxide dismutase strongly inhibited the loss of SH groups, indicating that H2O2 is involved in their oxidation. Fragmentation of BSA was observed during exposure to the XO + EDTA-Fe3+ and ascorbate + EDTA-Fe3+ systems and the products presented a broad band on sodium dodecyl sulfate polyacrylamide gel electrophoresis. Little formation of amine groups was observed in these systems, indicating that little peptide bond cleavage occurred. BSA exposed to the ascorbate + EDTA-Fe3+ system was more readily degraded by trypsin than that exposed to the XO + EDTA-Fe3+ system. Elastase degraded BSA exposed to the ascorbate + EDTA-Fe3+ system but not to the XO + EDTA-Fe3+ system.
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PMID:Oxidative damage to bovine serum albumin induced by hydroxyl radical generating systems of xanthine oxidase + EDTA-Fe3+ and ascorbate + EDTA-Fe3+. 133 12

A striking similarity exists between the pathogenetic properties of group A streptococci and those of activated mammalian professional phagocytes (neutrophils, macrophages). Both types of cells are endowed by the ability to adhere to target cells; to elaborate oxidants, hydrolases, and membrane-active agents (hemolysins, phospholipases); and to freely invade tissues and destroy cells. From the evolutionary point of view, streptococci might justifiably be considered the forefathers of "modern" leukocytes. Our earlier findings that synergy between a streptococcal hemolysin (streptolysin S, SLS) and a streptococcal thiol-dependent proteinase and between cytotoxic antibodies+complement and streptokinase-activated plasmin readily killed tumor cells, led us to hypothesize that by analogy to the pathogenetic mechanisms of streptococci, the mechanisms of tissue destruction initiated by activated leukocytes in inflammatory sites, as well as in tissues undergoing episodes of ischemia and reperfusion, might also be the result of the synergistic effects among leukocyte-derived oxidants, phospholipases, proteinases, cytokines, and cationic proteins. The current report extends our previous synergy studies with endothelial cells to two additional cell types--monkey kidney epithelial cells and rat beating heart cells. Monolayers of 51Cr-labeled cells that had been treated by combinations of sublytic amounts of hydrogen peroxide (generated either by glucose oxidase, xanthine-xanthine oxidase, or by paraquat) and with sublytic amounts of a variety of membrane-active agents (streptolysin S, phospholipases A2 and C, lysophosphatides, histone, chlorhexidine) were killed in a synergistic manner (double synergy). Crystalline trypsin markedly enhanced cell killing by combinations of oxidant and the membrane-active agents (triple synergy). Injury to the cells was characterized by the appearance of large membrane blebs that detached from the cells and floated freely in the media, looking like lipid droplets. Cytotoxicity induced by the various combinations of agonists was depressed, to a large extent, by scavengers of hydrogen peroxide (catalase, dimethyl thiourea, and by Mn2+) but not by SOD or by deferoxamine. When cationic agents were employed together with hydrogen peroxide, polyanions (heparin, polyanethole sulfonate) were also found to inhibit cell killing. It is proposed that in order to effectively combat the deleterious toxic effects of leukocyte-derived agonists on cells and tissues, antagonistic "cocktails" comprised of cationized catalase, cationized SOD, dimethylthiourea, Mn(2+)+glycine, proteinase inhibitors, putative inhibitors of phospholipases, and polyanions might be concocted. The current literature on synergistic phenomena pertaining to mechanisms of cell and tissue injury in inflammation is selectively reviewed.
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PMID:Synergism among oxidants, proteinases, phospholipases, microbial hemolysins, cationic proteins, and cytokines. 142 26

In vivo most extracellular iron is bound to transferrin or lactoferrin in such a way as to be unable to catalyze the formation of hydroxyl radical from superoxide (.O2-) and hydrogen peroxide (H2O2). At sites of Pseudomonas aeruginosa infection bacterial and neutrophil products could possibly modify transferrin and/or lactoferrin forming catalytic iron complexes. To examine this possibility, diferrictransferrin and diferriclactoferrin which had been incubated with pseudomonas elastase, pseudomonas alkaline protease, human neutrophil elastase, trypsin, or the myeloperoxidase product HOCl were added to a hypoxanthine/xanthine oxidase .O2-/H2O2 generating system. Hydroxyl radical formation was only detected with pseudomonas elastase treated diferrictransferrin and, to a much lesser extent, diferriclactoferrin. This effect was enhanced by the combination of pseudomonas elastase with other proteases, most prominently neutrophil elastase. Addition of pseudomonas elastase-treated diferrictransferrin to stimulated neutrophils also resulted in hydroxyl radical generation. Incubation of pseudomonas elastase with transferrin which had been selectively iron loaded at either the NH2- or COOH-terminal binding site yielded iron chelates with similar efficacy for hydroxyl radical catalysis. Pseudomonas elastase and HOCl treatment also decreased the ability of apotransferrin to inhibit hydroxyl radical formation by a Fe-NTA supplemented hypoxanthine/xanthine oxidase system. However, apotransferrin could be protected from the effects of HOCl if bicarbonate anion was present during the incubation. Apolactoferrin inhibition of hydroxyl radical generation was unaffected by any of the four proteases or HOCl. Alteration of transferrin by enzymes and oxidants present at sites of pseudomonas and other bacterial infections may increase the potential for local hydroxyl radical generation thereby contributing to tissue injury.
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PMID:Pseudomonas and neutrophil products modify transferrin and lactoferrin to create conditions that favor hydroxyl radical formation. 165 25

Activated human neutrophils (PMN) degrade rTNF-alpha resulting in a loss of cytotoxic activity against murine L-929 cells (L cells). This inactivation is mediated through proteases released from activated PMN. Exposure of TNF to H2O2, glucose oxidase, xanthine oxidase, or myeloper-oxidase-H2O2-halide did not affect TNF cytotoxicity for L cells. Exposure to trypsin, chymotrypsin, pronase E, or elastase, however, did diminish TNF bioactivity. FMLP-stimulated PMN in the presence, but not in the absence, of cytochalasin B reduced TNF activity, whereas PMA-stimulated PMN did not affect TNF. Stimulation of PMN with opsonized bacteria also induced TNF inactivation as well as the supernatant of FMLP-stimulated cells. Addition of protease inhibitors to the FMLP-stimulated cytochalasin B-treated PMN abrogated the inactivation of TNF cytotoxicity for L cells, whereas scavengers were not protective. In addition, PMN from a chronic granulomatous disease patient also decreased TNF bioactivity. Inactivation of TNF by activated PMN correlated with granule release and not with superoxide production. Exposure of TNF to proteases and FMLP-activated PMN also resulted in a loss of reactivity with anti-TNF antibodies, as measured by ELISA, and in the formation of an approximately 10-kDa split product from the 17-kDa rTNF molecule. Partial degradation of TNF by proteases released from activated PMN may result in a diminished TNF bioactivity and thereby contribute to the regulation of local inflammatory reactions.
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PMID:Inactivation of recombinant human tumor necrosis factor-alpha by proteolytic enzymes released from stimulated human neutrophils. 194 Mar 72

The role of intracellular oxyradicals in H2O2 and neutrophil-induced cytotoxicity is suggested by previous studies showing protection by inhibitors such as deferroxamine, dimethylthiourea, and dimethyl sulfoxide. In the current studies, the role of intracellular O2- is specifically examined by evaluating the effects of intracellular superoxide dismutase (SOD) supplementation on cytotoxicity of rat pulmonary artery endothelial cells induced by H2O2 and activated neutrophils. To minimize in vitro manipulation, supplementation was accomplished by incubating endothelial cells in the presence of SOD (1-20 mg/mL). Increases up to greater than 17-fold the baseline SOD activity were achievable using this approach, with uptake being maximal after 6 h of incubation. This increase was resistant to trypsin digestion, suggesting the intracellular location of SOD. Compared to controls, SOD-supplemented cells showed significantly increased resistance to killing by H2O2 and activated neutrophils. Inactive SOD failed to provide protection. The degree of protection was dependent on the dose of cytotoxic agent and the extent of SOD supplementation. The results provide new evidence that intracellular O2- participates in the killing process induced by these two stimuli. The intracellular source of O2- remains to be determined, although previous studies suggest xanthine oxidase as a likely candidate.
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PMID:Inhibition of cytotoxicity by intracellular superoxide dismutase supplementation. 212 22

The ability of various reactive oxygen species and serine proteases to activate latent collagenase (matrix metalloproteinase-1) purified from human neutrophils was examined. Latent 70-75 kD human neutrophil collagenase (HNC) was efficiently activated by known non-proteolytic activators phenylmercuric chloride (an organomercurial compound) and gold thioglucose (Au(I)-salt). Corresponding degree of activation was achieved by reactive oxygen species including hypochlorous acid (HOCl), hydrogen peroxide (H2O2) and hydroxyl radical generated by hypoxanthine/xanthine oxidase (HX/XAO). The presence of trace amounts of iron and EDTA were necessary and even enhanced H2O2 induced activation of latent HNC. This activation could be abolished by an iron chelator desferrioxamine and a hydroxyl radical scavenger mannitol. HOCl induced activation of latent HNC was not affected by desferrioxamine and mannitol. Thus, these compounds do not inhibit the active/activated form of HNC. Latent HNC could also be activated by trypsin and chymotrypsin but not by plasmin and plasma kallikrein. The ability of mannitol and desferrioxamine to inhibit the H2O2-induced activation of HNC suggests the transition metal dependent Fenton reaction to be responsible for localized and/or site-specific generation of hydroxyl radical/hydroxyl radical -like oxidants to act as the activating oxygen species. Our results support the ability of myeloperoxidase derived HOCl to act as a direct oxidative activator of HNC and further suggest the existence of a new/alternative oxidative activation pathway of HNC involving hydroxyl radical.
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PMID:Activation of latent human neutrophil collagenase by reactive oxygen species and serine proteases. 217 13


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