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:1.10.3.3 (ascorbate oxidase)
778 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Since cytochrome c and acetylated cytochrome c disappear from the circulation with a half-life of 4 min, these proteins cannot be used for in vivo detection of superoxide radicals and related metabolites. To determine superoxide and other radicals in vivo, a cytochrome c derivative (SMAC) was synthesized by linking 1 mol of poly(styrene-co-maleic acid) butyl ester (SM) to cytochrome c, followed by acetylation of its lysyl amino groups. SMAC retained 8 and 80% of cytochrome c activity to react with ascorbyl and superoxide radicals, respectively. However, SMAC did not serve as a substrate for cytochrome c reductase and cytochrome c oxidase. When injected intravenously to the rat, SMAC circulated bound to albumin with a half-life of 130 min. SMAC was rapidly reduced in the circulation of intact animals. Treatment of animals with paraquat markedly enhanced the reduction of the circulating SMAC. We have synthesized an SM-conjugated superoxide dismutase (SOD) derivative (SM-SOD) that circulates bound to albumin with a half-life of 6 h. Kinetic analysis revealed that SM-SOD effectively inhibited the superoxide-dependent reduction of SMAC either in the presence or absence of 0.5 mM albumin. However, the reduction of the circulating SMAC was not inhibited by SM-SOD both in normal and paraquat-treated animals. Plasma samples from both animal groups also reduced cytochrome c and SMAC by an SOD-insensitive mechanism. However, after treatment with ascorbate oxidase, both plasma samples lost their activity to reduce cytochrome c and SMAC. These and other results suggest that ascorbyl radical might principally be responsible for the reduction of circulating SMAC and that plasma levels of ascorbyl radical might increase in paraquat-treated animals.
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PMID:Synthesis of a cytochrome c derivative with prolonged in vivo half-life and determination of ascorbyl radicals in the circulation of the rat. 131 36

The reduction of ferricytochrome c within the perfusate in isolated lung perfusion systems has been demonstrated previously. We carried out the present study 1) to determine what reducing agents might be responsible for this reduction and 2) to determine whether the cytochrome c (cyto c) reduction within the recirculating perfusion system can be accounted for by relatively stable reducing agents released into the perfusate or whether some of the reduction is dependent on short-lived agents and/or proximity to the source of the agents within the lungs. Experiments were carried out with the use of isolated rabbit lungs perfused for 1 h in a recirculating system. In one group of experiments, ferricytochrome c was included in the recirculating perfusion system. In another group, the cyto c was added to produce the same concentration in samples after they were removed from a cyto c-free recirculating system. The recirculating cyto c was reduced at a rate of approximately 1.76 mumol/h, and approximately 22% was inhibitable by superoxide dismutase. Most of the rest could be inhibited by ascorbate oxidase within the recirculating perfusate. When the ferricytochrome c was added to the samples removed from the cyto c-free perfusion system, virtually the entire cyto c reducing capacity was inhibitable by ascorbate oxidase. Although reduced glutathione did accumulate in the recirculating perfusate, the quantity was not sufficient to have an important role in the cyto c reduction. We conclude that most of the cyto c reducing capacity within the lung perfusate could be accounted for by ascorbate released from the lungs.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Perfusate cytochrome c reduction in isolated rabbit lungs. 166 94

Enzymes and proteins: AO, amine oxidase; and as proposed in reference 3, BSAO, bovine serum AO; SSAO, swine serum AO; SKDAO, swine kidney AO; PSAO, pea seedling AO; APAO, arthrobacter P1AO; MADH, methylamine dehydrogenase; AAO, ascorbic acid oxidase; alpha-AE, alpha-amidating enzyme; Az, azurin; COX, cytochrome c oxidase; CP, ceruloplasmin; DBH, dopamine beta-hydroxylase; GO, galactose oxidase; Hc, hemocyanin; MT, metallotheonein; NIR, nitrite reductase; SOD, superoxide dismutase. Cofactors: Dopa, 3,4 dihydroxyphenylalanine; Topa, 3,4,6 trihydroxyphenyl-alanine; PLP, pyridoxal-phosphate; PQQ, pyrroloquinolinequinone. Reagents: DDC, diethyldithiocarbamate; DMG, diaminoguanidine; DMSA, dimercaptosuccinic acid; NTA, nitrilotriacetic acid. Technique-related: XANES, x-ray absorption near edge spectroscopy; EXAFS, extended x-ray absorption fine structure; ENDOR, electron-nuclear double resonance; ESEEM, electron spin echo envelope modulation; CD, circular dichroism; MCD, magnetic circular dichroism; NMRD, nuclear magnetic resonance dispersion; nqi, nuclear quadrupole interaction; DSC, differential scanning calorimetry.
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PMID:Copper in biological systems. A report from the 6th Manziana Conference, September 23-27, 1990. 175 86

Desferrioxamine (DFO) nearly doubles alkaline phosphatase oxidative inactivation by the ascorbate system. The effect is dependent on ascorbate and desferrioxamine concentrations, exhibiting in both cases a saturation mechanism. Conversion of desferrioxamine to ferrioxamine abolishes the prooxidant action. Desferrioxamine also increases ascorbate-dependent oxygen consumption and nitroblue tetrazolium reduction. Superoxide dismutase, which blocks the desferrioxamine enhancing effect on enzyme inactivation, markedly slows down nitroblue tetrazolium reduction as well as oxygen consumption by ascorbate plus desferrioxamine, while it fails to protect against the ascorbate system alone. Therefore, in the presence of desferrioxamine, the metal-catalyzed ascorbate autooxidation becomes superoxide-dependent and thus inhibitable by superoxide dismutase. Catalase, peroxidase, and ascorbate oxidase protect alkaline phosphatase from inactivation by both ascorbate and ascorbate-desferrioxamine systems. Hemin shields the enzyme from ascorbate plus DFO attack but not from ascorbate alone. In air-saturated solution, desferrioxamine seems to mediate one electron transfer from ascorbate to oxygen, generating superoxide anions, which can either trigger a Fenton reaction or produce desferal nitroxide radicals. In the absence of oxygen, ascorbate alone is ineffective, but the ascorbate plus desferrioxamine system still inactivates the enzyme; catalase, peroxidase, and ascorbate oxidase, but not superoxide dismutase, afford protection.
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PMID:Prooxidant action of desferrioxamine: enhancement of alkaline phosphatase inactivation by interaction with ascorbate system. 215 77

A method for the detection of ascorbate oxidase in electrophoretic gels is described. This method relies on the ability of the enzyme to prevent the photoreduction of nitroblue tetrazolium (NBT). The method is based on that described by C. Beauchamp and I. Fridovich (1971, Anal. Biochem. 44, 276-287) for the superoxide dismutase and was made specific for ascorbate oxidase detection by treating the gel with 0.1 M hydrogen peroxide. Ascorbate (25 microM) or riboflavin (500 microM) was used as the electron donor. The possible reaction mechanism in the presence of ascorbate has been investigated. Western and Northern blot analyses confirmed the results obtained from the NBT staining procedure.
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PMID:Electrophoretic detection of ascorbate oxidase activity by photoreduction of nitroblue tetrazolium. 222 47

The reactivity with nitric oxide was investigated for a number of type-1, type-2 and type-3 copper proteins azurin from Pseudomonas aeruginosa (type-1 copper); bovine superoxide dismutase, diamine oxidase from pig kidney and galactose oxidase from Dactylium dendroides (type-2 copper); haemocyanin from Helix pomatia (type-3 copper); the blue oxidases ceruloplasmin from pig serum, and ascorbate oxidase from Cucurbita pepo medullosa. Type-1 copper formed complexes with NO in the oxidised state, which complexes were only fully formed at low temperatures and could be photodissociated at 77K. Complex formation led to the disappearance of the EPR signal of type-1 copper and of the optical absorbance band in the 600 nm region. In azurin, photodissociation caused the reappearance of the original 625 nm absorbance band, but in the blue oxidases, a new band with lower intensity was found at 595 nm instead of the original absorbance band at 610 nm. In all cases, the EPR signal of type-1 copper did not return. These results are best explained by the formation of a photolabile type-1 Cu1+-NO+ complex. They also indicate that in the complex formed, the type-1 copper structure is probably not disrupted, and that after illumination, the nitric oxide molecule is still in the near vicinity of the copper atom. Type-2 copper did not react at all with nitric oxide, and type-3 copper formed complexes with nitric oxide in both the oxidised and the reduced state, but photodissociation of these complexes could not be demonstrated.
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PMID:The reaction of nitric oxide with copper proteins and the photodissociation of copper-NO complexes. 282 26

Experiments were carried out to evaluate the production of hydroxyl radical-like species by intact rat liver cells by assaying for the production of ethylene from alpha-keto-4-thiomethylbutyric acid in the absence and presence of added iron. In the absence of iron, a low rate of ethylene production, which was not sensitive to superoxide dismutase, catalase, or competitive scavengers was observed. Ethylene was produced when KMBA was incubated with perfusates of rat liver or the suspension medium obtained after incubating liver cells for varying periods of time, followed by removal of the liver cells. Boiling the perfusate or suspension medium had no effect on ethylene production. This ethylene production does not appear to reflect an oxygen radical-mediated event. The addition of ferric-EDTA, but not ferric-desferrioxamine, increased ethylene production by the hepatocyte suspensions in a reaction sensitive to inhibition by catalase, ascorbate oxidase, and competitive scavengers, but not superoxide dismutase. The sensitivity to externally added catalase and ascorbate oxidase suggested that the ethylene production reflected an extracellular oxygen radical generating system. Ferric alone and several ferric chelates, for example, ferric-ATP, ADP, AMP, histidine, and citrate stimulated ethylene production using perfusates of liver or suspension medium after removal of the hepatocytes. The sensitivity of the added iron system to ascorbate oxidase suggested that during perfusion or incubation of liver cells, efflux of ascorbate occurs, followed by reduction of the iron and subsequently, extracellular production of oxygen radicals.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Ethylene production from alpha-keto-4-thiomethylbutyric acid by isolated rat liver cells, suspension medium, and perfusates in the absence and presence of iron. 301 75

Tyrosinase usually catalyzes the conversion of monophenols to o-diphenols and oxidation of diphenols to the corresponding quinones. However, when 3,4-dihydroxymandelic acid was provided as the substrate, it catalyzed an unusual oxidative decarboxylation reaction generating 3,4-dihydroxybenzaldehyde as the sole product. The identity of the product was confirmed by high-performance liquid chromatography (HPLC) as well as ultraviolet and infrared spectral studies. None of the following enzymes tested catalyzed the new reaction: galactose oxidase, ceruloplasmin, superoxide dismutase, ascorbate oxidase, dopamine beta-hydroxylase, and peroxidase. Phenol oxidase inhibitors such as phenylthiourea, potassium cyanide, and sodium azide inhibited the reaction drastically, suggesting the participation of the active site copper of the enzyme in the catalysis. Mimosine, a well-known competitive inhibitor of tyrosinase, competitively inhibited the new reaction also. 4-Hydroxymandelic acid and 3-methoxy-4-hydroxymandelic acid neither served as substrates nor inhibited the reaction. Putative intermediates such as 3,4-dihydroxybenzyl alcohol and (3,4-dihydroxybenzoyl)formic acid did not accumulate during the reaction. Oxidation to a quinone methide derivative rather than conventional quinone accounts for this unusual oxidative decarboxylation reaction. Earlier from this laboratory, we reported the conversion of 4-alkylcatechols to quinone methides catalyzed by a cuticular phenol oxidase [Sugumaran, M., & Lipke, H. (1983) FEBS Lett. 155, 65-68]. Present studies demonstrate that mushroom tyrosinase will also catalyze quinone methide production with the same active site copper if a suitable substrate such as 3,4-dihydroxymandelic acid is provided.
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PMID:Tyrosinase catalyzes an unusual oxidative decarboxylation of 3,4-dihydroxymandelate. 309 74

The presence of ascorbate free radical (AFR) reductase (NADH:AFR oxidoreductase, EC 1.6.5.4) in senile cataractous human lenses was demonstrated by measuring spectrophotometrically NADH oxidation in the presence of ascorbate plus ascorbate oxidase. About 80-85% of the lens AFR reductase was probably recovered in the supernatant of the lens homogenate. Michaelis constants of the reductase were about 10 microM and less than 1 microM for AFR and NADH, respectively. We also showed that AFR reductase activities in the cataractous lenses tended to decrease with increase of insoluble lens protein contents, or showed rather the possibility that the reductase activity may have decreased before the lens protein aggregation. In the highest activity group (about 120-160 nmol NADH oxidized/min/lens), it was roughly calculated that the reductase in the lens could re-reduce immediately the total (or almost total) amount of AFR produced there by ascorbate oxidation even at a high rate of 600-800 microM/min, if NADH concentration in the lens were sufficiently maintained. The above results suggested that AFR reductase in the human lens plays important roles in ascorbate regeneration of its redox cycle, and that activity loss of AFR reductase, as well as of superoxide dismutase, glutathione peroxidase and glutathione reductase, may be responsible for the oxidative changes in lens proteins with the development of senile cataracts.
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PMID:Ascorbate free radical reductase and ascorbate redox cycle in the human lens. 318 51

The dietary antagonism between copper and molybdate salts prompted a study of the inhibition of copper enzymes by thiomolybdate (TM). TM strongly inhibited the oxidase activity of five copper oxidase with I50% values in the 1-5 microM range. The mechanism of the TM effect on the copper oxidase, ceruloplasmin (Cp) (E.C. 1.16.3.1), was studied in detail. In Vmax vs. E plots, TM gave parallel data suggesting irreversibility but a large number of TM molecules per Cp were required. The inhibition of Cp by TM could not be reversed by dialysis. Isolation of TM-inhibited Cp on Sephadex G-10 did not yield any active Cp molecules. Cu(II) did not restore any inhibited oxidase activity. Gel electrophoresis supported the covalent binding of Cp by TM without any extensive change in protein structure. EPR results confirmed that Cu(II) is reduced to Cu(I) after reaction with TM. However, the Mo(VI) in MoS4(2-) did not change in oxidation number. Analysis of the TM-Cp compound accounted for all six Cu atoms as found in native Cp. The data suggest the covalent binding of sulfide to Cp copper. TM also inhibited the activity of ascorbate oxidase, cytochrome oxidase, superoxide dismutase, and tyrosinase. However, no inhibition of carbonic anhydrase, a zinc enzyme, was observed at 1 mM TM.
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PMID:Inhibition of ceruloplasmin and other copper oxidases by thiomolybdate. 609 47


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