Nitrite reduction to nitrous oxide by propionibacteria: Detoxication mechanism

Characteristics of dissimilatory nitrate reduction by Propionibacterium acidi-propionici, P. freudenreichii, P. jensenii, P. shermanii and P. thoenii were studied. All strains reduced nitrate to nitrite and further to N₂O. Recovery of added nitrite-N as N₂O-N approached 100%, so that no other end product existed in a significant quantity. Specific rates of N₂O production were 3 to 6 orders of magnitude lower than specific rates of N₂ production by common denitrifiers. Oxygen but not acetylene inhibited N₂O production in P. acidi-propionici and P. thoenii. Nitrite reduction rates were generally higher than nitrate reduction rates. The enzymes involved in nitrate and nitrite reduction were either constitutive or derepressed by anacrobiosis. Nitrate stimulated synthesis of nitrate reductase in P. acidi-propionici. Specific growth rates and growth yields were increased by nitrate. At 10 mM, nitrite was toxic to all strains, and at 1 mM its effect ranged from none to total inhibition. No distinction was obvious between incomplete forms of denitrification and dissimilatory nitrate reduction to ammonia. N₂O production from nitrite by propionibacteria may represent a detoxication mechanism rather than a part of an energy transformation system.     

The effect of chloride on nitrate reductase level,on anaerobic nitrite production,and on nitrate content in excisedPisum sativum L. roots

The effect was studied of chloride ions, added in the form of different salts, on nitrate reductase (NR) level in excised pea roots, on anaerobic nitrite production in an assay medium lacking both nitrate and n-propanol, on nitrate content in the roots, and on in vivo NR activity determined in an assay medium containing 5% n-propanol. The presence of Cl in nitrate containing nutrient solutions resulted in lower NR levels, however counterions supplied together with Cl tended to modify slightly this general trend. The negative effect of Cl ions was also apparent, when Cl ions were applied before nitrate ions. Anaerobic nitrite production in the medium lacking both nitrate and n-propanol was not influenced by chloride ions. Nitrate content in the roots was reduced in the presence of chloride both at 3 mM and 15 mM NO₃ in nutrient solutions; however, at 16 mM NO₃, nitrate content in the roots exoeeded even in the presence of 15 mM Cl nitrate content in those root segments which were cultivated in a nutrient solution with 6 mM nitrate, which is the concentration at which NR reaches the level of saturation in excised pea roots. The results obtained suggest that a special induction nitrate pool exists in plant cells besides the storage and metabolic nitrate pools.     

Aerobic nitrate and nitrite reduction in continuous cultures of Escherichia coli E4

Nitrate and nitrite was reduced by Escherichia coli E4 in a l-lactate (5 mM) limited culture in a chemostat operated at dissolved oxygen concentrations corresponding to 90–100% air saturation. Nitrate reductase and nitrite reductase activity was regulated by the growth rate, and oxygen and nitrate concentrations. At a low growth rate (0.11 h^–1) nitrate and nitrite reductase activities of 200 nmol · mg^–1 protein · min^–1 and 250 nmol · mg^–1 protein · min^–1 were measured, respectively. At a high growth rate (0.55 h^–1) both enzyme activities were considerably lower (25 and 12 nmol mg^–1 · protein · min^–1). The steady state nitrite concentration in the chemostat was controlled by the combined action of the nitrate and nitrite reductase. Both nitrate and nitrite reductase activity were inversely proportional to the growth rate. The nitrite reductase activity decreased faster with growth rate than the nitrate reductase. The chemostat biomass concentration of E. coli E4, with ammonium either solely or combined with nitrate as a source of nitrogen, remained constant throughout all growth rates and was not affected by nitrite concentrations. Contrary to batch, E. coli E4 was able to grow in continuous cultures on nitrate as the sole source of nitrogen. When cultivated with nitrate as the sole source of nitrogen the chemostat biomass concentration is related to the activity of nitrate and nitrite reductase and hence, inversely proportional to growth rate.     

Aluminium effect on nitrate assimilation in cucumber (Cucumis sativus L.) roots

In vivo effect of aluminium on nitrate uptake and reduction by cucumber seedlings was investigated. The high-performance liquid chromatography was used to analyse the rate of nitrate uptake. Low (0.5 mM) concentration of AlCl₃ in the nutrient solution stimulated nitrate uptake during the first 3 h. On the other hand, 6 h exposure of the cucumber seedlings to 1 or 5 mM of AlCl₃ resulted in inhibition of nitrate uptake and at 5 mM concentration of AlCl₃ the efflux of nitrate was observed. Furthermore, the amount of nitrate accumulated in cucumber roots after aluminium treatment was decreased. The noteworthy fact was observed, that at all concentrations of aluminium tested on increase of the nitrate reductase activity. This stimulation was concentration depended, but independent of the source of the enzyme. The activity of both the cytosolic and the plasma membrane bound nitrate reductase activity was enhanced in vivo. On the other hand, AlCl₃ applied in vitro only slighty decreased nitrate reductase activity.     

Effect of aerobic and anaerobic conditions on the in vivo nitrate reductase assay in spinach leaves

¹⁵N-labelled nitrate was used to show that nitrate reduction by leaf discs in darkness was suppressed by oxygen, whereas nitrite present within the cell could be reduced under aerobic dark conditions. In other experiments, unlabelled nitrite, allowed to accumulate in the tissue during the dark anaerobic reduction of nitrate was shown by chemical analysis to be metabolised during a subsequent dark aerobic period. Leaves of intact plants resembled incubated leaf discs in accumulating nitrite under anaerobic conditions. Nitrate, n-propanol and several respiratory inhibitors or uncouplers partly reversed the inhibitory effect of oxygen on nitrate reduction in leaf discs in the dark. Of these nitrate and propanol acted synergistically. Reversal was usually associated with inhibition of respiration but some concentrations of 2,4-dinitrophenol (DNP) and ioxynil reversed inhibition without affecting respiratory rates. Respiratory inhibitors and uncouplers stimulated nitrate reduction in the anaerobic in vivo assay i.e. in conditions where the respiratory process is non-functional. Freezing and thawing leaf discs diminished but did not eliminate the sensitivity of nitrate reduction to oxygen inhibition.Abbreviations DNP 2,4-dinitrophenol - HOQNO 8-hydroxyquinoline-N-oxide - DCPIP 2,6-dichlorophenolindophenol - CCCP Carbonyl cyanide m-chlorophenylhydrazone - TES N-tris(hydroxymethyl)methyl-2-amino ethanesulphonic acid - HEPES N-2-hydroxyethylpiperazine-N-2-ethanesulphonic acid     

nir1, a conditional-lethal mutation in barley causing a defect in nitrite reduction

Summary Eleven green individuals were isolated when 95000 M₂ plants of barley (Hordeum vulgare L.), mutagenised with azide in the M₁, were screened for nitrite accumulation in their leaves after nitrate treatment in the light. The selected plants were maintained in aerated liquid culture solution containing glutamine as sole nitrogen source. Not all plants survived to flowering and some others that did were not fertile. One of the selected plants, STA3999, from the cultivar Tweed could be crossed to the wild-type cultivar and analysis of the F₂ progeny showed that leaf nitrite accumulation was due to a recessive mutation in a single nuclear gene, which has been designated Nir1. The homozygous nir1 mutant could be maintained to flowering in liquid culture with either glutamine or ammonium as sole nitrogen source, but died within 14 days after transfer to compost. The nitrite reductase cross-reacting material seen in nitrate-treated wild-type plants could not be detected in either the leaf or the root of the homozygous nir1 mutant. Nitrite reductase activity, measured with dithionite-reduced methyl viologen as electron donor, of the nitrate-treated homozygous nir1 mutant was much reduced but NADH-nitrate reductase activity was elevated compared to wild-type plants. We conclude that the Nir1 locus determines the formation of nitrite reductase apoprotein in both the leaf and root of barley and speculate that it represents either the nitrite reductase apoprotein gene locus or, less likely, a regulatory locus whose product is required for the synthesis of nitrite reductase, but not nitrate reductase. Elevation of NADH-nitrate reductase activity in the nir1 mutant suggests a regulatory perturbation in the expression of the Narl gene.     

Anaerobic expression of nitric oxide reductase from denitrifying Pseudomonas stutzeri

NO reductase synthesis was investigated immunochemically and by activity assays in cells of Pseudomonas stutzeri ZoBell grown in continuous culture at discrete aeration levels, or in O₂-limited batch cultures supplemented with N oxides as respiratory substrate. Under aerobic conditions, NO reductase was not expressed in P. stutzeri. Oxygen limitation in combination with the presence of nitrate or nitrite derepressed NO reductase synthesis. On transition from aerobic to anaerobic conditions in continuous culture, NO reductase was synthesized below 3% air saturation and reached maximum expression under anaerobic conditions. By use of mutant strains defective in nitrate respiration or nitrite respiration, the inducing effect of individual N oxides on NO reductase synthesis could be discriminated. Nitrite caused definite, concentration-dependent induction, while nitrate promoted moderate enzyme synthesis or amplified effects of nitrite. Exogenous nitric oxide (NO) in concentrations 25 M induced trace amounts of NO reductase; in higher concentrations it arrested cell growth. Nitrite reductase or NO reductase were not detected immunochemically under these conditions. NO generated as an intermediate appeared not to induce NO reductase significantly. Antiserum raised against the P. stutzeri NO reductase showed crossreaction with cell extracts from P. stutzeri JM300, but not with several other denitrifying pseudomonads or Paracoccus denitrificans.     

Control of nitrate and nitrite assimilation by carbohydrate reserves,adenosine nucleotides and pyridine nucleotides in leaves of Zea mays L. under dark conditions

The rate of in-vivo nitrate reduction by leaf segments of Zea mays L. was found to decline during the second hour of dark anaerobic treatment. On transfer to oxygen the capacity to reduce nitrate under dark conditions was restored. These observations led to the proposal that nitrate reductase is a regulatory enzyme with ADP acting as a negative effector. The effect of ADP on the invitro activity of nitrate reductase and the changes in the in-vivo adenylate pool under dark-N₂ and dark-O₂ were investigated. It was found that ADP inhibited the activity of partially purified nitrate reductase. Similarly, the in-vivo anaerobic inhibition of nitrate reduction was associated with a build-up of ADP in the leaf tissue. Under anaerobic conditions nitrite accumulated and on transfer to oxygen the accumulated nitrite was reduced. To explain this phenomenon the following hypothesis was proposed and tested. Under anaerobic conditions the supply of reducing equivalents for nitrite reduction in the plastid becomes restricted and nitrite accumulates as a consequence. On transfer to oxygen this restriction is removed and nitrite disappears. This capacity to reduce accumulated nitrite was found to be dependent on the carbohydrate status of the leaf tissue.     

The influence of pretreatment with different cations on anaerobic nitrite production by excisedPisum sativum roots

The influence of pretreatment with some cations on anaerobic nitrite production (in an assay medium lacking nitrate) by excised primary roots of pea (Pisum sativum L., ov. Raman), detached from six-day-old seedlings germinated in distilled water, was investigated. When the excised roots were precultivated in one-salt-solutions of KNO3, then these roots produced at 9 mM and 15 mM NO3- concentrations under anaerobic conditions significantly more NO2-, than those precultivated in a nutrient solution containing besides K+ ions also Ca2+ and Mg2+ ions, and they produced nitrite for a longer time. The KNO3 dependent increase in anaerobic NO2- production was counteracted most by Ca2+ and to a lesser extent by Mg2+; Na+ was without effect. NH4+ at higher concentrations (12 and 15 mM) significantly depressed nitrite production both by roots precultivated in a solution containing besides NH4+ only K+, and by roots precultivated in a full nutrient solution containing K+, Ca2+ and Mg2+, however at lower NH4+ concentrations (0.6 and 2mMNH4+; 15mMNO3-) the decrease was more conspicuous in the KNO3 solution than in the full nutrient solution. Nitrate reductase level was not influenced by this pretreatment. When 6% and 7.5% n-propanol, which increases membrane permeability and causes mixing of storage and metabolic nitrate pools in the cells, was added to the assay medium lacking nitrate, anaerobic nitrite production increased and the differences caused by the precultivation disappeared. These results show that higher K+ concentrations in unbalanced one-salt-solutions of KNO3 can cause higher membrane permeability by accentuating Ca8+ deficiency, which results in a faster penetration of NO3- from the storage pool to the sites of its reduction and in an easier penetration of NO2- out of the roots, and that higher NH4+ concentrations can change nitrate compartmentation and diminish the metabolic NO3- pool which results in a slower nitrate reduction. Besides that, lower NH4+ concentrations in KNO3 solutions (15mMNO3-) probably partially counteract the K+ dependent increase in membrane permeability. The results obtained show that there is no simple, direct relationship between the so-called metabolic pool of nitrate (i.e. anaerobic nitrite production) and the level of nitrate reductase, but that the velocity of nitrate reduction can be influenced by nitrate compartmentation in the cell.     

Differential effect of tungsten on the development of endogenous and nitrate-induced nitrate reductase activities in soybean leaves

The effect of tungsten on the development of endogenous and nitrate-induced NADH- and FMNH₂-linked nitrate reductase activities in primary leaves of 10-day-old soybean (Glycine max [L.] Merr.) seedlings was studied. The seedlings were grown with or without exogenous nitrate. High levels of endogenous nitrate reductase activities developed in leaves of seedlings grown without nitrate. However, no endogenous nitrite reductase activity was detected in such seedlings. The FMNH₂-linked nitrate reductase activity was about 40% of NADH-linked activity. Tungsten had little or no effect on the development of endogenous NADH- and FMNH₂-linked nitrate reductase activities, respectively. By contrast, in nitrate-grown seedlings, tungsten only inhibited the nitrate-induced portion of NADH-linked nitrate reductase activity, whereas the FMNH₂-linked activity was inhibited completely. Tungsten had no effect on the development of nitrate-induced nitrite reductase activity. The complete inhibition of FMNH₂-linked nitrate reductase activity by tungsten in nitrate-grown plants was apparently an artifact caused by the reduction of nitrite by nitrite reductase in the assay system. The results suggest that in soybean leaves either the endogenous nitrate reductase does not require molybdenum or the molybdenum present in the seed is preferentially utilized by the enzyme complex as compared to nitrate-induced nitrate reductase.     

A heme-C-containing enzyme complex that exhibits nitrate and nitrite reductase activity from the dissimilatory iron-reducing bacterium Geobacter metallireducens

Nitrate reduction in the dissimilatory iron-reducing bacterium Geobacter metallireducens was investigated. Nitrate reductase and nitrite reductase activities in nitrate-grown cells were detected only in the membrane fraction. The apparent K _m values for nitrate and nitrite were determined to be 32 and 10 μM, respectively. Growth on nitrate was not inhibited by either tungstate or molybdate at concentrations of 1 mM or less, but was inhibited by both at 10 and 20 mM. Nitrate and nitrite reductase activity in the membrane fraction was not, however, affected by dialysis with 20 mM tungstate. An enzyme complex that exhibited both nitrate and nitrite reductase activity was solubilized from membrane fractions with CHAPS and was partially purified by preparative gel electrophoresis. It was found to be composed of four different polypeptides with molecular masses of 62, 52, 36, and 16 kDa. The 62-kDa polypeptide [a low-midpoint potential (–207 mV), multiheme cytochrome c] exhibited nitrite reductase activity under denaturing conditions. No molybdenum was detected in the complex by plasma-emission mass spectrometry. Received: 26 March 1999 / Accepted: 16 August 1999     

Nitrate and Nitrite Reduction in Relation to Nitrogenase Activity in Soybean Nodules and Rhizobium japonicum Bacteroids

Soybean (Glycine max L. cv Williams) seeds were sown in pots containing a 1:1 perlite-vermiculite mixture and grown under greenhouse conditions. Nodules were initiated with a nitrate reductase expressing strain of Rhizobium japonicum, USDA 110, or with nitrate reductase nonexpressing mutants (NR⁻ 108, NR⁻ 303) derived from USDA 110. Nodules initiated with either type of strain were normal in appearance and demonstrated nitrogenase activity (acetylene reduction). The in vivo nitrate reductase activity of N₂-grown nodules initiated with nitrate reductase-negative mutant strains was less than 10% of the activity shown by nodules initiated with the wild-type strain. Regardless of the bacterial strain used for inoculation, the nodule cytosol and the cell-free extracts of the leaves contained both nitrate reductase and nitrite reductase activities. The wild-type bacteroids contained nitrate reductase but not nitrite reductase activity while the bacteroids of strains NR⁻ 108 and NR⁻ 303 contained neither nitrate reductase nor nitrite reductase activities.

Addition of 20 millimolar KNO₃ to bacteroids of the wild-type strain caused a decrease in nitrogenase activity by more than 50%, but the nitrate reductase-negative strains were insensitive to nitrate. The nitrogenase activity of detached nodules initiated with the nitrate reductase-negative mutant strains was less affected by the KNO₃ treatment as compared to the wild-type strain; however, the results were less conclusive than those obtained with the isolated bacteroids.

The addition of either KNO₃ or KNO₂ to detached nodules (wild type) suspended in a semisolid agar nutrient medium caused an inhibition of nitrogenase activity of 50% and 65% as compared to the minus N controls, and provided direct evidence for a localized effect of nitrate and nitrite at the nodule level. Addition of 0.1 millimolar sucrose stimulated nitrogenase activity in the presence or absence of nitrate or nitrite. The sucrose treatment also helped to decrease the level of nitrite accumulated within the nodules.

     

Semiaerobic induction of a nitrate reductase activity in growing cultures of Rhodopseudomonas capsulata E1F1 containing NH 4 + ions as nitrogen source

Rhodopseudomonas capsulata E₁F₁ growing under chemo- or photoorganotrophic conditions shows nitrate reductase activity which:

1. Is not repressed by ammonium ions;
2. Is governed by the partial pressure of oxygen in the gas phase.

Upon induction of nitrate reductase activity under optimum conditions, and also in the presence of amonium ions, subsequent addition of nitrate does not result in nitrite production. The specific activity of the enzyme increases by a factor of 10, when the growth medium is supplemented by the addition of nitrate. The physiological role of this ammonium resistant activity is not known.     

Isolation and characterization of a chlorate-resistant mutant (Clo- R) of the symbiotic cyanobacterium Nostoc ANTH: heterocyst formation and N(2)-fixation in the presence of nitrate,and evidence for separate nitrate and nitrite transport systems

Nostoc ANTH is a filamentous, heterocystous cyanobacterium capable of N₂-fixation in the absence of combined nitrogen. A chlorate-resistant mutant (Clo-R) of Nostoc ANTH was isolated that differentiates heterocysts and fixes N₂ in the presence of nitrate, but not in the presence of nitrite or ammonium. The mutant lacks nitrate uptake and thereby also lacks induction of nitrate reductase activity by nitrate. However, this mutant is able to transport and assimilate nitrite, indicating that there is a transport system for nitrite that is distinct from that for the nitrate. The lack of inhibitory effect of nitrate on N₂-fixation was owing to lack of nitrate uptake and not to lack of enzymes for its assimilation (nitrate reductase and glutamine synthetase) or the lack of an ammonium transport system for retention of ammonia. The mutant has potential for use as a biofertilizer supplementing chemical nitrate fertilizer in rice fields, without N₂-fixation being adversely affected. Received: 16 October 2001 / Accepted: 26 November 2001     

Utilization of nitrate by bacteroids of Bradyrhizobium japonicum in the soybean root nodule

Bacteroids of Bradyrhizobium japonicum strain CB1809, unlike CC705, do not have a high level of constitutive nitrate reductase (NR; EC 1.7.99.4) in the soybean (Glycine max. Merr.) nodule. Ex planta both strains have a high activity of NR when cultured on 5 mM nitrate at 2% O₂ (v/v). Nitrite reductase (NiR) was active in cultured cells of bradyrhizobia, but activity with succinate as electron donor was not detected in freshly-isolated bacteroids. A low activity was measured with reduced methyl viologen. When bacteroids of CC705 were incubated with nitrate there was a rapid production of nitrite which resulted in repression of NR. Subsequently when NiR was induced, nitrite was utilized and NR activity recovered. Nitrate reductase was induced in bacteroids of strain CB1809 when they were incubated in-vitro with nitrate or nitrite. Increase in NR activity was prevented by rifampicin (10 g· ml^-1) or chloramphenicol (50 g·ml^-1). Nitrite-reductase activity in bacteroids of strain CB1809 was induced in parallel with NR. When nitrate was supplied to soybeans nodulated with strain CC705, nitrite was detected in nodule extracts prepared in aqueous media and it accumulated during storage (1°C) and on further incubation at 25°C. Nitrite was not detected in nodule extracts prepared in ethanol. Thus nitrite accumulation in nodule tissue appears to occur only after maceration and although bacteroids of some strains of B. japonicum have a high level of a constitutive NR, they do not appear to reduce nitrate in the nodule because this anion does not gain access to the bacteroid zone. Soybeans nodulated with strains CC705 and CB1809 were equally sensitive to nitrate inhibition of N₂ fixation.Abbreviations NR nitrate reductase - NiR nitrite reductase - Tris 2-amino-2-(hydroxymethyl)-1,3-propanediol     

Nitrate reductase-deficient mutant cell lines of Nicotiana tabacum

Summary The wild-type line and 14 nitrate reductase-deficient mutant cell lines of Nicotiana tabacum were tested for the presence of nitrate reductase partial activities, and for nitrite reductase and xanthine dehydrogenase activity. Data characterizing the electron donor specificity of nitrate reductase (EC 1.6.6.1., NADH:nitrate oxidoreductase) and nitrite reductase (EC 1.7.7.1., ferredoxin:nitrite oxidoreductase) of the wild-type line are presented. Three lines (designated cnx) simultaneously lack NADH-, FADH₂-, red. benzyl viologen-nitrate reductase, and xanthine dehydrogenase activities, but retain the nitrate reductase-associated NADH-cytochrome c reductase activity. These mutants are, therefore, interpreted to be impaired in gene functions essential for the synthesis of an active molybdenum-containing cofactor. For cnx-68 and cnx-101, the sedimentation coefficient of the defective nitrate reductase molecules does not differ from that of the wild-type enzyme (7.6S). In 11 lines (designated nia) xanthine dehydrogenase activity is unaffected, and the loss of NADH-nitrate reductase is accompanied by a loss of all partial activities, including NADH-cytochrome c reductase. However, one line (nia-95) was found to possess a partially active nitrate reductase molecule, retaining its FADH₂- and red. benzyl viologen nitrate reductase activity. It is likely that nia-95 is a mutation in the structural gene for the apoprotein. Both, the nia and cnx mutant lines exhibit nitrite reductase activity, being either nitrate-inducible or constitutive. Evidence is presented that, in Nicotiana tabacum, nitrate, without being reduced to nitrite, is an inducer of the nitrate assimilation pathway.     

A comparison of the influence of iron on the growth and nitrate metabolism of Anabaena and Scenedesmus

A comparative study of growth and nitrate metabolism of Anabaena flos-aquae (Lyng.) Bréb. and Scenedesmus bijugatus var. seriatus Chodat investigated possible mechanisms for the iron-stimulated increases in growth specific for blue-green algae in mixed algal communities. Algae were separately grown in an morganic medium with varying concentrations of iron and nitrate to determine the effects on each organism. Iron was found to be a limiting nutrient for cultures of both Anabaena and Scenedesmus as determined by chlorophyll a concentrations and cell enumeration. Both iron and nitrate stimulated the specific activity of nitrate reductase, nitrite reductase, and glutamine synthetase in Anabaena. Iron enrichment did not increase the activity of the enzymes in Scenedesmus, but inhibited the activity of nitrate reductase and glutamine synthetase. The stimulation of growth by iron in cells grown under iron limiting conditions was associated with increased nitrate metabolism in Anabaena but not in Scenedesmus.     

The effect of oxygen on nitrate and nitrite assimilation in leaves of Zea mays L. under dark conditions

The assimilation of nitrate under dark-N₂ and dark-O₂ conditions in Zea mays leaf tissue was investigated using colourimetric and ¹⁵N techniques for the determination of organic and inorganic nitrogen. Studies using ¹⁵N indicated that nitrate was assimilated under dark conditions. However, the rate of nitrate assimilation in the dark was only 28% of the rate under non-saturating light conditions. No nitrite accumulated under dark aerobiosis, even though nitrate reduction occurred under these conditions. The pattern of nitrite accumulation in leaf tissue in response to dark-N₂ conditions consisted of three phases: an initial lag phase, followed by a period of rapid nitrite accumulation and finally a phase during which the rate of nitrite accumulation declined. After a 1-h period of dark-anaerobiosis, both nitrate reduction and nitrite accumulation declined considerably. However, when O₂ was supplied, nitrate reduction was stimulated and the accumulated nitrite was rapidly reduced. Anaerobic conditions stimulated nitrate reduction in leaf tissue after a period of dark-aerobic pretreatment.     

Mutants of Azotobacter vinelandii altered in the regulation of nitrate assimilation

The two enzymes involved in the assimilatory pathway of nitrate in Azotobacter vinelandii are corregulated. Nitrate reductase and nitrite reductase are inducible by nitrate and nitrite. Ammonium represses induction by nitrate of both reductases. Repression by ammonium is higher in media containing 2-oxo-glutarate as carbon source than in media containing sucrose. Mutants in the gene ntrC lost nitrate and nitrite reductase simultaneously. Ten chlorate-resistant mutants with a new phenotype were isolated. In media without ammonium they had a normal phenotype, being sensitive to the toxic effect of chlorate. In media containing low ammonium concentrations they were resistant to chlorate. These mutants seem to be affected in the repression of nitrate and nitrite reductases by ammonium.