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1.
On transition from nitrogen starvation to ammonia or ammonia/glutamine sufficiency Phormidium uncinatum produces high amounts of H2O2, which is consumed by several oxidative reactions catalyzed by thylakoid membrane bound enzymes. These include: oxidation of glutamine to free hydroxylamine, of ammonia to nitrite, of bound hydroxylamine to nitrite, and dismutation of free hydroxylamine to ammonia and nitrite. A possible role of these transformations for detoxification is discussed.Non-standard abbreviations FCCP
p-trifluormethoxy carbonylcyanide phenylhydrazone
- DCMU
dichloromethyl urea 相似文献
2.
Intact cells of the denitrifying fungus Fusarium oxysporum were previously shown to catalyze codenitrification to form a hybrid nitrous oxide (N2O) species from nitrite and other nitrogen compounds such as azide and ammonia. Here we show that cytochrome P450nor can catalyze the codenitrification reaction to form N2O from nitric oxide (NO) but not nitrite, and azide or ammonia. The results show that the direct substrate of the codenitrification by intact cells should not be nitrite but NO, which is formed from nitrite by the reaction of a dissimilatory nitrite reductase. 相似文献
3.
Nitrite reductases found in plants, algae, and cyanobacteria catalyze the six-electron reduction of nitrite to ammonia with reduced ferredoxin serving as the electron donor. They contain one siroheme and one [4Fe-4S] cluster, acting as separate one-electron carriers. Nitrite is thought to bind to the siroheme and to remain bound until its complete reduction to ammonia. In the present work the enzyme catalytic cycle, with ferredoxin reduced by photosystem 1 as an electron donor, has been studied by EPR and laser flash absorption spectroscopy. Substrate depletion during enzyme turnover, driven by a series of laser flashes, has been demonstrated. A complex of ferrous siroheme with NO, formed by two-electron reduction of the enzyme complex with nitrite, has been shown to be an intermediate in the enzyme catalytic cycle. The same complex can be formed by incubation of free oxidized nitrite reductase with an excess of nitrite and ascorbate. Hydroxylamine, another putative intermediate in the reduction of nitrite catalyzed by nitrite reductase, was found to react with oxidized nitrite reductase to produce the same ferrous siroheme-NO complex, with a characteristic formation time of about 13 min. The rate-limiting step for this reaction is probably hydroxylamine binding to the enzyme, with the conversion of hydroxylamine to NO at the enzyme active site likely being much faster. 相似文献
4.
Rocco L. Mancinelli Sonja Cronin Lawrence I. Hochstein 《Archives of microbiology》1986,145(2):202-208
Paracoccus halodenitrificans, grown anaerobically in the presence of nitrite, contained membrane and cytoplasmic nitrite reductases. When assayed in the presence of phenazine methosulfate and ascorbate, the membranebound enzyme produced nitrous oxide whereas the cytoplasmic enzyme produced nitric oxide. When both enzymes were assayed in the presence of methyl viologen and dithionite, the cytoplasmic enzyme produced ammonia. Following solubilization, the membrane-bound enzyme behaved like the cytoplasmic enzyme, producing nitric oxide in the presence of phenazine methosulfate and ascorbate, and ammonia when assayed in the presence of methyl viologen and dithionite. The cytoplasmic and membranebound enzymes were purified to essentially the same specific activity. Only a single nitrite-reductase activity was detected on electrophoretic gels and the electrophoretic behavior of both enzymes suggested they were identical. The spectral properties of both enzymes suggested they were cd-type cytochromes. These data suggest that the products of nitrite reduction by the cd-cytochrome nitrite reductase are determined by the location of the enzyme and the redox potential of the electron donor.Abbreviations PMS
phenazine methosulfate
- MV
methyl viologen
- HEPES
N-2-hydroxyethylpiperazine-N-2-ethane-sulfonic acid
- CHAPSO
[3-(3-cholamidopropyldimethylammonia)-1-(2-hydroxy-1-propanesulfonate)]
National Research Council Research Fellow 相似文献
5.
Nitrite is a breakdown product of nitric oxide that in turn is oxidized to nitrate in cells. In this work, we investigated whether reactive oxidant species might be generated during nitrite metabolism in cultured EA.hy926 endothelial cells. Nitrite was taken up by the cells in a time- and concentration-dependent manner and oxidized to nitrate, which accumulated in cells to concentrations almost 10-fold those of nitrite. Conversion of low millimolar concentrations of nitrite to nitrate was associated with increased oxidant stress in the cells. This manifested as increased oxidation of dihydrofluorescein in tandem with depletion of both GSH and ascorbate. Further, loading cells with ascorbate or treatment with desferrioxamine prevented nitrite-induced dihydrofluorescein oxidation. Nitrite within cells also increased the fluorescence of 4-amino-5-methylamino-2′,7′-difluorofluorescein and inhibited the activity of cellular glyceraldehyde 3-phosphate dehydrogenase, which are markers of intracellular nitrosation reactions. Intracellular ascorbate partially prevented both of these effects of nitrite. Although ascorbate can reduce nitrite to nitric oxide at low pH, in endothelial cells loaded with ascorbate, its predominant effect at high nitrite concentrations is to prevent potentially damaging nitrosation reactions. 相似文献
6.
Nitrite is a breakdown product of nitric oxide that in turn is oxidized to nitrate in cells. In this work, we investigated whether reactive oxidant species might be generated during nitrite metabolism in cultured EA.hy926 endothelial cells. Nitrite was taken up by the cells in a time- and concentration-dependent manner and oxidized to nitrate, which accumulated in cells to concentrations almost 10-fold those of nitrite. Conversion of low millimolar concentrations of nitrite to nitrate was associated with increased oxidant stress in the cells. This manifested as increased oxidation of dihydrofluorescein in tandem with depletion of both GSH and ascorbate. Further, loading cells with ascorbate or treatment with desferrioxamine prevented nitrite-induced dihydrofluorescein oxidation. Nitrite within cells also increased the fluorescence of 4-amino-5-methylamino-2',7'-difluorofluorescein and inhibited the activity of cellular glyceraldehyde 3-phosphate dehydrogenase, which are markers of intracellular nitrosation reactions. Intracellular ascorbate partially prevented both of these effects of nitrite. Although ascorbate can reduce nitrite to nitric oxide at low pH, in endothelial cells loaded with ascorbate, its predominant effect at high nitrite concentrations is to prevent potentially damaging nitrosation reactions. 相似文献
7.
Nitrate Reduction to Nitrite,Nitric Oxide and Ammonia by Gut Bacteria under Physiological Conditions
The biological nitrogen cycle involves step-wise reduction of nitrogen oxides to ammonium salts and oxidation of ammonia back to nitrites and nitrates by plants and bacteria. Neither process has been thought to have relevance to mammalian physiology; however in recent years the salivary bacterial reduction of nitrate to nitrite has been recognized as an important metabolic conversion in humans. Several enteric bacteria have also shown the ability of catalytic reduction of nitrate to ammonia via nitrite during dissimilatory respiration; however, the importance of this pathway in bacterial species colonizing the human intestine has been little studied. We measured nitrite, nitric oxide (NO) and ammonia formation in cultures of Escherichia coli, Lactobacillus and Bifidobacterium species grown at different sodium nitrate concentrations and oxygen levels. We found that the presence of 5 mM nitrate provided a growth benefit and induced both nitrite and ammonia generation in E.coli and L.plantarum bacteria grown at oxygen concentrations compatible with the content in the gastrointestinal tract. Nitrite and ammonia accumulated in the growth medium when at least 2.5 mM nitrate was present. Time-course curves suggest that nitrate is first converted to nitrite and subsequently to ammonia. Strains of L.rhamnosus, L.acidophilus and B.longum infantis grown with nitrate produced minor changes in nitrite or ammonia levels in the cultures. However, when supplied with exogenous nitrite, NO gas was readily produced independently of added nitrate. Bacterial production of lactic acid causes medium acidification that in turn generates NO by non-enzymatic nitrite reduction. In contrast, nitrite was converted to NO by E.coli cultures even at neutral pH. We suggest that the bacterial nitrate reduction to ammonia, as well as the related NO formation in the gut, could be an important aspect of the overall mammalian nitrate/nitrite/NO metabolism and is yet another way in which the microbiome links diet and health. 相似文献
8.
An analysis of the genome sequence database revealed novel types of two-domain multi-copper oxidases. The two-domain proteins have the conspicuous combination of blue-copper and inter-domain trinuclear copper binding residues, which is common in ceruloplasmin and ascorbate oxidase but not in nitrite reductase, and therefore are considered to retain the characteristics of the plausible ancestral form of ceruloplasmin and ascorbate oxidase. A possible evolutionary relationship of these proteins is proposed. 相似文献
9.
Nitrite reductase (EC 1.6.6.4) has been purified 730-fold from spinach leaves. The enzyme catalyzes the reduction of nitrite to ammonia, with the use of reduced form of methyl viologen and ferredoxin. A stoichiometry of one molecule of nitrite reduced per molecule of ammonia formed has been found. KCN at 2.5×10-4 m inhibited nitrite reductase activity almost completely. Purified enzyme was almost homogeneous by disk electrophoresis with polyacrylamide gel. The molecular weight of the enzyme was estimated to be 61,000 from gel filtration. Nitrite reductase, in the oxidized form, has absorption maxima at 276, 388 and 573 mμ. Both methyl viologen and ferredoxin linked nitrite reductase activities of the enzyme were inactivated on exposure to low ionic strength. 相似文献
10.
Purification of a hexaheme cytochrome c552 from Escherichia coli K 12 and its properties as a nitrite reductase 总被引:2,自引:0,他引:2
Anaerobic cytochrome c552 was purified to electrophoretic homogeneity by ion-exchange chromatography and gel filtration from a mutant of Escherichia coli K 12 that synthesizes an increased amount of this pigment. Several molecular and enzymatic properties of the cytochrome were investigated. Its relative molecular mass was determined to be 69 000 by sodium dodecyl sulfate/polyacrylamide gel electrophoresis. It was found to be an acidic protein that existed in the monomeric form in the native state. From its heme and iron contents, it was concluded to be a hexaheme protein containing six moles of heme c/mole protein. The amino-acid composition and other properties of the purified cytochrome c552 indicated its similarity to Desulfovibrio desulfuricans hexaheme cytochrome. The cytochrome c552 showed nitrite and hydroxylamine reductase activities with benzyl viologen as an artificial electron donor. It catalyzed the reduction of nitrite to ammonia in a six-electron transfer. FMN and FAD also served as electron donors for the nitrite reduction. The apparent Michaelis constants for nitrite and hydroxylamine were 110 microM and 18 mM, respectively. The nitrite reductase activity of the cytochrome c552 was inhibited effectively by cupric ion and cyanide. 相似文献
11.
Hendrikus J. Laanbroek Marie-Jos Br-Gilissen Hans L. Hoogveld 《Applied microbiology》2002,68(3):1454-1457
Ammonia-starved cells of Nitrosomonas europaea are able to preserve a high level of ammonia-oxidizing activity in the absence of ammonium. However, when the nitrite-oxidizing cells that form part of the natural nitrifying community do not keep pace with the ammonia-oxidizing cells, nitrite accumulates and may subsequently inhibit ammonia oxidation. The maintenance of a high ammonia-oxidizing capacity during starvation is then nullified. In this study we demonstrated that cells of N. europaea starved for ammonia were not sensitive to nitrite, either when they were starved in the presence of nitrite or when nitrite was supplied simultaneously with fresh ammonium. In the latter case, the initial ammonia-oxidizing activity of starved cells was stimulated at least fivefold. 相似文献
12.
Summary
Anabaena cylindrica was immobilized in calcium alginate beads and was placed in a batch reactor in the presence of a glutamine synthetase inhibitor (methionine sulfoximide). Ammonia was released in the medium during two days with a rate of 0.35 moles h–1mgChl–1. Addition of nitrite to the medium increased the ammonia production as cells used the nitrite reductase pathway to form ammonia. When reactors were placed in anaerobiosis by N2 bubbling, ammonia production was sustained several days and the total ammonia formed was about two fold higher than in aerobiosis. Long term effects of MSX, nitrite and anaerobiosis are discussed. 相似文献
13.
Variable Ammonia Production Among Smooth and Rough Strains of Pseudomonas pseudomallei: Resemblance to Bacteriocin Production 总被引:4,自引:1,他引:3
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The colonial morphology of some strains of Pseudomonas pseudomallei was correlated with certain biochemical and physiological traits. After 3 days of growth on Wahba or heart infusion agars, smooth-colony strains generated toxic amounts of ammonia. Under the same conditions, the rough strains simultaneously produced oxalic acid which decreased the inhibitory concentration of ammonia. The ammonia-ammonium concentrations in smooth cultures exhibited certain bacteriocin-like characteristics. An unusually stable, smooth strain (strain 165) was chosen to compare and emphasize any differences with typical, rough strain 7815. Three-day-old smooth cultures grown on Wahba agar containing 3% (w/v) glycerol demonstrated ammonia toxicity. The substitution of glucose for glycerol completely obviated this toxicity. In highly aerated Wahba broth containing glucose, the amount of ammonia found in strain 165 smooth cultures and the amount of oxalic acid found in strain 7815 rough cultures were greatly reduced. In Difco nitrate broth smooth strain 165 did not form gas, and it reduced nitrate to nitrite only. Strain 7815 produced a gas and reduced both nitrate and nitrite. 相似文献
14.
Nitric oxide, when released into the bloodstream, is quicklyscavenged by Hb in erythrocytes or oxidized to nitrite. Nitrite canalso enter erythrocytes and oxidize Hb. The goals of this work were todetermine the mechanism of erythrocyte nitrite uptake and whether thisuptake causes oxidant stress in these cells. Erythrocytes took up 0.8 mM nitrite with a half-time of 11 min. Nitrite uptake was sensitive totemperature and to the pH and ionic composition of the medium but wasnot inhibited by the specific anion-exchange inhibitor DIDS. About 25%of nitrite uptake occurred on the sodium-dependent phosphatetransporter and the rest as diffusion of nitrous acid or other speciesacross the plasma membrane. Methemoglobin formation increased inproportion to the intracellular nitrite concentration. Nitritereacted with erythrocyte ascorbate, but ascorbate loading of cellsdecreased nitrite-induced methemoglobin formation only at high nitriteconcentrations. In conclusion, nitrite rapidly enters erythrocytes andreacts with oxyhemoglobin but does not exert a strong oxidant stress onthese cells. 相似文献
15.
Tripathy JN Hirasawa M Kim SK Setterdahl AT Allen JP Knaff DB 《Photosynthesis research》2007,94(1):1-12
A system has been developed for expressing a His-tagged form of the ferredoxin-dependent nitrite reductase of spinach in Escherichia coli. The catalytic and spectral properties of the His-tagged, recombinant enzyme are similar, but not identical, to those previously
observed for nitrite reductase isolated directly from spinach leaf. A detailed comparison of the spectral, catalytic and fluorescence
properties of nitrite reductase variants, in which each of the enzyme’s eight tryptophan residues has been replaced using
site-directed mutagenesis by either aromatic or non-aromatic amino acids, has been used to examine possible roles for tryptophan
residues in the reduction of nitrite to ammonia catalyzed by the enzyme. 相似文献
16.
Ammonia oxidation by the methane oxidising bacterium Methylococcus capsulatus strain bath 总被引:1,自引:0,他引:1
Soluble extracts of Methylococcus capsulatus (Bath) that readily oxidise methane to methanol will also oxidise ammonia to nitrite via hydroxylamine. The ammonia oxidising activity requires O2, NADH and is readily inhibited by methane and specific inhibitors of methane mono-oxygenase activity. Hydroxylamine is oxidised to nitrite via an enzyme system that uses phenazine methosulphate (PMS) as an electron acceptor. The estimated K
mvalue for the ammonia hydroxylase activity was 87 mM but the kinetics of the oxidation were complex and may involve negative cooperativity.Abbreviations PMS
Phenazine methosulphate
- NADH
nicotinamide adenine dinucleotide, reduced form
-
K
m
Michaelis constant
- NO
2
-
nitrite
- NH2OH
hydroxylamine 相似文献
17.
The effect of different levels of salt, sodium nitrite, polyphosphate and various sugars on growth, pigment production, protease activity and culture pH caused by Monascus purpureus was studied in broth medium and ground meat. The addition of sodium chloride (> 50.0 g l(-1)) and polyphosphate (> 3.0g l(-1)) to broth medium decreased mycelial growth, pigment production and protease activity of M. purpureus, whereas low concentrations of sodium nitrite (< 0.2 g l(-1)) promoted mycelial growth and pigment production. When the basal medium and ground meat contained salt, 150.0 g l(-1), the mould growth was stopped. The medium with fructose as carbon source proved to be the most suitable for mycelium growth and pigment production, with maltose and glucose being the second most productive. When sucrose and lactose were used as carbon sources, mycelium growth and pigment production were inhibited but the protease activity increased significantly. The mould showed more tolerance to salt and polyphosphate in ground meat than in broth medium and used sucrose as a carbon source as well as glucose for growth and pigment production in the meat mixture. 相似文献
18.
N-acetyl-l -glutamine, pyroglutamic acid, and the butyl ester of pyroglutamic acid were isolated in pure form from an aqueous extract of human brain. These compounds were isolated by combination of paper and ion exchange chromatography. The isolated substance identified as N-acetyl-l -glutamine did not react with the ninhydrin reagent but yielded glutamic acid and ammonia upon acid hydrolysis. An acetyl hydrazide was identified by paper chromatography from hydrazinolysates of the isolated substance. The glutamic acid liberated by hydrolysis had the l -configuration. The results of elementary analysis of the isolated compound were in full accord with the analysis calculated for synthetic N-acetyl-l -glutamine. A large amount of pyroglutamic acid and a substance identical with the butyl ester of pyroglutamic acid were isolated in pure form. The results of our studies suggest that pyroglutamic and the butyl ester derivative were artifacts formed during the isolation and purification procedures. 相似文献
19.
Mikhail Beketov 《Hydrobiologia》2004,528(1-3):209-216
Complex toxic effects of ammonia, nitrite and nitrate to aquatic animals are not well investigated. In rivers of southwestern Siberia, Russia, elevated ammonia and nitrite concentrations corresponded to significant reduction in species diversity of mayflies. The objectives of the study were to evaluate the sensitivity of six mayfly species to the mixture of ammonia, nitrite and nitrate in acute laboratory tests and to compare the sensitivity found with results of the first river bioassessment in the region considered and with species saprobic indexes. The rank of the species sensitivity was: Baetis vernus < Potamanthus luteus < B. fuscatus=Cloeon bifidum < Ephemerella lenoki < Heptagenia sulphurea (p < 0.05). The experiments revealed variation in sensitivity among the species by a factor of 7.5. Comparison of the tests results and the available field data shows that species which exhibit higher tolerance in the tests inhabit comparatively greater amount of sites include contaminated places. Final conclusion is addressed in comparison of the results found with more spatially and temporally extensive observations. Saprobic indexes of the species and their acute tolerances (LC50s) tend to be positively correlated (r=0.93, p=0.02). 相似文献
20.
Nitrite Uptake into Intact Pea Chloroplasts : II. Influence of Electron Transport Regulators, Uncouplers, ATPase and Anion Uptake Inhibitors and Protein Binding Reagents 总被引:6,自引:3,他引:3
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The relationship between net nitrite uptake and its reduction in intact pea chloroplasts was investigated employing electron transport regulators, uncouplers, and photophosphorylation inhibitors. Observations confirmed the dependence of nitrite uptake on stromal pH and nitrite reduction but also suggested a partial dependance upon PSI phosphorylation. It was also suggested that ammonia stimulates nitrogen assimilation in the dark by association with stromal protons. Inhibition of nitrite uptake by N-ethylmaleimide and dinitrofluorobenzene could not be completely attributed to their inhibition of carbon dioxide fixation. Other protein binding reagents which inhibited photosynthesis showed no effect on nitrite uptake, except for p-chlormercuribenzoate which stimulated nitrite uptake. The results with N-ethylmaleimide and dinitrofluorobenzene tended to support the proposed presence of a protein permeation channel for nitrite uptake in addition to HNO2 penetration. On the basis of a lack of effect by known anion uptake inhibitors, it was concluded that the nitrite uptake mechanism was distinct from that of phosphate and chloride/sulfate transport. 相似文献