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)

As part of an ongoing study of the role of subcellular fractions on the metabolism of nitroxides, we studied the metabolism of a set of five nitroxides in cytosol derived from rat hepatocytes. The nitroxides were chosen to provide information on the effects of the type of charge and the ring on which the nitroxyl group is located. The rates of reduction were fastest for a six-membered positively charged nitroxide ('CAT-1') and slowest for an anionic five-membered ring nitroxide ('PCA'). Changing levels of glutathione, sulphydryl groups in general, NADPH or NADH had little or no effect on the rates of reduction, while the addition of ascorbate oxidase essentially abolished reduction of the nitroxides. The products of reduction by the cytosol were the corresponding hydroxylamines. The overall rates of reduction of neutral or anionic nitroxides were much slower than those observed with intact cells. We conclude that the primary source of metabolism of nitroxides by cytosol is reduction by ascorbate and that under most conditions reduction of nitroxides in the cytosol is not a major factor in the metabolism of nitroxides by cells.
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PMID:Metabolism of nitroxide spin labels in subcellular fractions of rat liver. II. Reduction in the cytosol. 236 85

Differences in oxidative metabolism between subsarcolemmal and interfibrillar heart mitochondria were investigated. Interfibrillar mitochondria oxidized substrates donating reducing equivalents at Complex I (NADH-CoQ reductase), Complex II (succinate-CoQ reductase), and Complex III (CoQH2-cytochrome c reductase) more rapidly than did subsarcolemmal mitochondria. There was no difference in oxidation of substrates entering the electron transport chain at Complex IV (cytochrome c oxidase). Differences expressed in normal-ionic-strength medium at Complexes II and III but not I were eliminated in low-ionic-strength medium. The concentrations of cytochromes and activities of NADH and cytochrome c oxidase were virtually the same in the two populations. In permeabilized mitochondria, activities of succinate-duroquinone and TMPD plus ascorbate oxidase were significantly lower in the subsarcolemmal mitochondria. Differences in membrane permeability between the populations were suggested by the greater permeability of subsarcolemmal mitochondria to exogenous NADH. The influence of isolation buffers and preparative procedures on the two classes of mitochondria were also examined. Characteristic biochemical and morphological properties of the two populations were unchanged by exposing each to the preparative procedure used to isolate the alternate population; the oxidative performance of the two populations cannot be equalized by experimental manipulation.
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PMID:Biochemical differences between subsarcolemmal and interfibrillar mitochondria from rat cardiac muscle: effects of procedural manipulations. 298 22

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

Coated vesicles were isolated from rat liver in about 80% fraction purity as determined from electron microscopy and analyses of marker enzymes and compared with Golgi apparatus and other membrane fractions isolated in parallel. The fractions were enriched in NADH-monodehydroascorbate reductase, ascorbate oxidase and ascorbic acid. The NADH-monodehydroascorbate reductase and ascorbate oxidase of the Golgi apparatus and coated vesicles differed from that of the endoplasmic reticulum in being inhibited by the sodium selective ionophore, monensin, at physiological concentrations while these activities were stimulated by ethylenediaminetetraacetic acid in coated vesicles but not in Golgi apparatus. Activities of both coated vesicles and Golgi apparatus fractions depleted in the coat protein, clathrin, were activated by the addition of clathrin-rich supernatant fractions. The results are discussed in the context of monodehydroascorbate as an acceptor for electron transport-mediated transfer of electrons from NADH by coated vesicles as part of a possible mechanism to drive membrane translocations or to acidify the interiors of vesicles.
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PMID:Monodehydroascorbate as an electron acceptor for NADH reduction by coated vesicle and Golgi apparatus fractions of rat liver. 614 8

The inhibitory effect of organic cations from the group of hydrazones on phosphorylated respiration in mitochondria with a succinate as a substrate has been shown to be completely reversed by 2,4-dinitrophenol (DNP). The effect observed has not been due to the action directly on the ATP-synthetase because the inhibition of ATPase, NADH-succinate oxidase and ascorbate oxidase activities occurs at a wide concentration range of compounds studied. The concentration of organic cations used for the inhibition of the respiration in submitochondria particles (SMP) as compared with respiration in mitochondria is higher; in case of SMP the inhibitory effect is not reversed by DNP. These results allow to conclude that the high inhibitory effect of organic cations is due to the energy-dependent binding of these compounds to the enzyme in the membrane phase of mitochondria.
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PMID:[Energy-dependent inhibition of mitochondrial NAD.H-oxidase and succinic oxidase by strong nitrous bases]. 661 29

The formation of ascorbate radicals, identified by ESR experiments, was observed in the ascorbate peroxidase reaction by thyroid microsomes. The steady-state concentration of ascorbate radicals decreased in the presence of NADH. The oxidation of NADH was followed optically. Using the ascorbic acid oxidase system, NADH-dependent electron transport in thyroid microsomes was examined. Ascorbate radicals competed with bound cytochrome b5 for the reaction with reduced NADH-cytochrome b5 reductase. The NADH-ascorbate radical reductase activity of thyroid microsomes was calculated to be 0.17 mumol/mg.s at 3.3 microM ascorbate radicals. Kinetic results show that the properties of NADH-cytochrome b5 reductase in thyroid microsomes were similar to those of the enzyme in liver microsomes. The formation of ascorbate radicals by thyroid microsomes was stimulated by the addition of thyroxine, and the stimulation was decreased also by NADH. The thyroxine-mediated oxidation of ascorbate is explained in terms of consecutive one-electron transfers initiated by bound thyroid peroxidase. These results, along with those described in our previous paper (M. Nakamura, I. Yamazaki, and S. Ohtaki, 1990, J. Biochem. 108, 804-810), support the idea that ascorbate protects thyroid hormones from oxidative degradation through the NADH-cytochrome b5 reductase system.
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PMID:Formation and reduction of ascorbate radicals by hog thyroid microsomes. 839 46

Recycling of ascorbic acid from its oxidized forms is required to maintain intracellular stores of the vitamin in most cells. Since the ubiquitous selenoenzyme thioredoxin reductase can recycle dehydroascorbic acid to ascorbate, we investigated the possibility that the enzyme can also reduce the one-electron-oxidized ascorbyl free radical to ascorbate. Purified rat liver thioredoxin reductase catalyzed the disappearance of NADPH in the presence of low micromolar concentrations of the ascorbyl free radical that were generated from ascorbate by ascorbate oxidase, and this effect was markedly stimulated by selenocystine. Dehydroascorbic acid is generated by dismutation of the ascorbyl free radical, and thioredoxin reductase can reduce dehydroascorbic acid to ascorbate. However, control studies showed that the amounts of dehydroascorbic acid generated under the assay conditions used were too low to account for the observed loss of NADPH. Electron paramagnetic resonance spectroscopy directly confirmed that the reductase decreased steady-state ascorbyl free radical concentrations, as expected if thioredoxin reductase reduces the ascorbyl free radical. Dialyzed cytosol from rat liver homogenates also catalyzed NADPH-dependent reduction of the ascorbyl free radical. Specificity for thioredoxin reductase was indicated by loss of activity in dialyzed cytosol prepared from livers of selenium-deficient rats, by inhibition with aurothioglucose at concentrations selective for thioredoxin reductase, and by stimulation with selenocystine. Microsomal fractions prepared from rat liver showed substantial NADH-dependent ascorbyl free radical reduction that was not sensitive to selenium depletion. These results suggest that thioredoxin reductase can function as a cytosolic ascorbyl free radical reductase that may complement cellular ascorbate recycling by membrane-bound NADH-dependent reductases.
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PMID:Reduction of the ascorbyl free radical to ascorbate by thioredoxin reductase. 972 29

This paper describes the development of a modified electrode for the electrocatalytic oxidation of beta-nicotinamide adenine dinucleotide (beta-NADH) and beta-nicotinamide adenine dinucleotide phosphate (beta-NADPH) using electropolymerised 3,4-dihydroxybenzaldehyde (3,4-DHB). Two voltammetric biosensors using enzyme-immobilised membranes were constructed for the determination of formic acid and glucose-6-phosphate (G6P), respectively. The formic acid biosensor based on the combination of formate dehydrogenase (FDH)-modified membrane with 3,4-DHB-coated glassy carbon electrode is one to two orders more sensitive (LOD, 5.0x10(-5) M) than previously reported electrochemical biosensors. Similarly, lower detection limit (4.0x10(-5) M) for the measurement of G6P was achieved using glucose-6-phosphate dehydrogenase (G6PDH) in the presence of beta-NADP(+). The interference of uric acid and ascorbate was minimised by incorporating an additional membrane modified with uricase and ascorbate oxidase, respectively. The biosensing scheme developed in this study can be adopted universally with a number of dehydrogenases for the detection of different substrates.
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PMID:Voltammetric biosensors for the determination of formate and glucose-6-phosphate based on the measurement of dehydrogenase-generated NADH and NADPH. 1134

Reduction of the ascorbate free radical (AFR) at the plasma membrane provides an efficient mechanism to preserve the vitamin in a location where it can recycle alpha-tocopherol and thus prevent lipid peroxidation. Erythrocyte ghost membranes have been shown to oxidize NADH in the presence of the AFR. We report that this activity derives from an AFR reductase because it spares ascorbate from oxidation by ascorbate oxidase, and because ghost membranes decrease steady-state concentrations of the AFR in a protein- and NADH-dependent manner. The AFR reductase has a high apparent affinity for both NADH and the AFR (< 2 microM). When measured in open ghosts, the reductase is comprised of an inner membrane activity (both substrate sites on the cytosolic membrane face) and a trans-membrane activity that mediates extracellular AFR reduction using intracellular NADH. However, the trans-membrane activity constitutes only about 12% of the total measured in ghosts. Ghost AFR reductase activity can also be differentiated from NADH-dependent ferricyanide reductase(s) by its sensitivity to the detergent Triton X-100 and insensitivity to enzymatic digestion with cathepsin D. This NADH-dependent AFR reductase could serve to recycle ascorbic acid at a crucial site on the inner face of the plasma membrane.
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PMID:Recycling of the ascorbate free radical by human erythrocyte membranes. 1142 97

Plasma membrane vesicles from adult rat brain synaptosomes (PMV) have an ascorbate-dependent NADH oxidase activity of 35-40 nmol/min/(mg protein) at saturation by NADH. NADPH is a much less efficient substrate of this oxidase activity, with a Vmax 10-fold lower than that measured for NADH. Ascorbate-dependent NADH oxidase activity accounts for more than 90% of the total NADH oxidase activity of PMV and, in the absence of NADH and in the presence of 1 mm ascorbate, PMV produce ascorbate free radical (AFR) at a rate of 4.0 +/- 0.5 nmol AFR/min/(mg protein). NADH-dependent *O2- production by PMV occurs with a rate of 35 +/- 3 nmol/min/(mg protein), and is a coreaction product of the NADH oxidase activity, because: (i) it is inhibited by more than 90% by addition of ascorbate oxidase, (ii) it is inhibited by 1 micro g/mL wheat germ agglutinin (a potent inhibitor of the plasma membrane AFR reductase activity), and (iii) the KM(NADH) of the plasma membrane NADH oxidase activity and of NADH-dependent *O2- production are identical. Treatment of PMV with repetitive micromolar ONOO- pulses produced almost complete inhibition of the ascorbate-dependent NADH oxidase and *O2- production, and at 50% inhibition addition of coenzyme Q10 almost completely reverts this inhibition. Cytochrome c stimulated 2.5-fold the plasma membrane NADH oxidase, and pretreatment of PMV with repetitive 10 microm ONOO- pulses lowers the K0.5 for cytochrome c stimulation from 6 +/- 1 (control) to 1.5 +/- 0.5 microm. Thus, the ascorbate-dependent plasma membrane NADH oxidase activity can act as a source of neuronal.O2-, which is up-regulated by cytosolic cytochrome c and down-regulated under chronic oxidative stress conditions producing ONOO-.
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PMID:The NADH oxidase activity of the plasma membrane of synaptosomes is a major source of superoxide anion and is inhibited by peroxynitrite. 1215 84


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