Superoxide production during reduction of molecular oxygen by assimilatory nitrate reductase
| Autor: | Christopher J. Kay, Michael J. Barber |
|---|---|
| Rok vydání: | 1996 |
| Předmět: |
Cyanide
Biophysics Chlorella Nitrate reductase Photochemistry Biochemistry Adduct Superoxide dismutase chemistry.chemical_compound Nitrate Nitrate Reductases Superoxides Molecular Biology chemistry.chemical_classification biology Chemistry Superoxide Nitrate Reductase (NAD(P)H) Osmolar Concentration Electron acceptor Hydrogen-Ion Concentration Oxygen Kinetics Catalase biology.protein Oxidation-Reduction Nuclear chemistry |
| Zdroj: | Archives of biochemistry and biophysics. 326(2) |
| ISSN: | 0003-9861 |
| Popis: | Assimilatory NADH:nitrate reductase catalyzes the transfer of reducing equivalents from NADH to molecular oxygen. Initial rate studies performed under conditions of optimum pH (8.0) and constant ionic strength (μ = 0.2) revealed that the maximal rate of activity with molecular oxygen was 0.5% (0.044 μmol NADH consumed/min/nmol heme) that of the activity using[formula]as the terminal electron acceptor (9.0 μmol NADH consumed/min/nmol heme) with a K m for O 2 of 586 μ M . NADH:molecular oxygen reductase activity exhibited a pH optimum of 9.2, was inhibited by cyanide, and was unaffected by changes in ionic strength or the presence of phosphate ions. Spectroscopic studies indicated NADH:molecular oxygen reductase activity resulted in the production of the superoxide radical, detected as the formation of adrenochrome from epinephrine and by the reduction of nitroblue tetrazolium, both of which could be inhibited by the addition of superoxide dismutase and were unaffected by the addition of catalase. Direct observation of superoxide production using spin-trapping in combination with EPR spectroscopy resulted in the detection of the spin adduct 5,5-dimethyl-5-hydroxy-1-pyrrolidinyloxy (DMPO-OH). The formation of this spin adduct was abolished either in the absence of nitrate reductase, NADH, or DMPO or by the addition of superoxide dismutase or nitrate and was greatly reduced by the presence of cyanide. Inclusion of catalase or ethanol had no effect on the formation of the spin adduct. These results indicate that nitrate reductase can utilize molecular oxygen as an electron acceptor and that the product,[formula]is primarily generated via the Mo-pterin center. |
| Databáze: | OpenAIRE |
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