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Query: EC:1.7.1.2 (nitrate reductase)
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The genes encoding the structural components of the nitrate/nitrite assimilation system of the oceanic cyanobacterium Synechococcus sp. strain WH 8103 were cloned and characterized. The genes encoding nitrate reductase (narB) and nitrite reductase (nirA) are clustered on the chromosome but are organized in separate transcriptional units. Upstream of narB is a homologue of nrtP that encodes a nitrate/nitrite-bispecific permease rather than the components of an ABC-type nitrate transporter found in freshwater cyanobacteria. Unusually, neither nirA nor ntcA (encoding a positive transcription factor of genes subject to nitrogen control) were found to be tightly regulated by ammonium. Furthermore, transcription of glnA (encoding glutamine synthetase) is up-regulated in ammonium-grown cells, highlighting significant differences in nitrogen control in this cyanobacterium. Nitrogen depletion led to the transient up-regulation of ntcA, nirA, nrtP, narB, and glnA in what appears to be an NtcA-dependent manner. The NtcA-like promoters found upstream of nirA, nrtP, and narB all differ in sequence from the canonical NtcA promoter established for other cyanobacteria, and in the case of nirA, the NtcA-like promoter was functional only in cells deprived of combined nitrogen. The ecological implications of these findings are discussed in the context of the oligotrophic nature of oceanic surface waters in which Synechococcus spp. thrive.
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PMID:Nitrate/nitrite assimilation system of the marine picoplanktonic cyanobacterium Synechococcus sp. strain WH 8103: effect of nitrogen source and availability on gene expression. 1466 Mar 43

Nitrogen metabolism is not only one of the basic processes of plant physiology, but also one of the important parts of global chemical cycle. Plant nitrogen assimilation directly takes part in the synthesis and conversion of amino acid through the reduction of nitrate. During this stage, some key enzymes, e.g., nitrate reductase (NR), glutamine synthetase (GS), glutamate dehydrogenase (GDH), glutamine synthase (GOGAT), aspargine synthetase (AS), and asparate aminotransferase (AspAT) participate these processes. The protein is assimilated in plant cell through amino acid, and becomes a part of plant organism through modifying, classifying, transporting and storing processes, etc. The nitrogen metabolism is associated with carbonic metabolism through key enzyme regulations and the conversion of products, which consists of basic life process. Among these amino acids in plant cell, glutamic acid (Glu), glutamine (Gln), aspartic acid (Asp) and asparagines (Asn), etc., play a key role, which regulates their conversion each other and their contents in the plant cell through regulating formation and activity of those key enzymes. Environmental factors also affect the conversion and recycle of the key amino acids through regulating gene expression of the key enzymes and their activities. Nitrate and light intensity positively regulate the gene transcription of NR, but ammonium ions and Glu, Gln do the negative way. Water deficit is a very serious constraint on N2 fixation rate and soybean (Glycine max Merr.) grain yield, in which, ureide accumulation and degradation under water deficit appear to be the key issues of feedback mechanism on nitrogen fixation. Water stress decreases NR activity, but increases proteinase activity, and thus, they regulate plant nitrogen metabolism, although there are some different effects among species and cultivars. Water stress also decreases plant tissue protein content, ratio of protein and amino acid, and reduces the absorption of amino acid by plant. On the contrary, soil flooding decreases the content and accumulation amount of root nitrogen in winter wheat by 11.9% from booting to flowering stages and 39.1% during grain filling stage, and reduces the ratio of carbon and nitrogen by 79.6%. The results misadjust the metabolism between carbon and nitrogen, and result in the end of the root growth. Elevated CO2 level could decrease plant leaf nitrogen content under well-watered condition, but almost maintain stable under water deficit condition. The radiation of UV-B significantly reduces the partitioning coefficient and synthetic rate of Rubisco, which significantly decreases the photosynthetic rate. This paper reviewed the pathway of plant nitrogen assimilation, characteristics of key enzymes and their regulating mechanisms with picturing the regulating mode of NR, and described the signal sensing and conduct of plant nitrogen metabolism and the formation, transportation, storage and degradation of plant cell protein with picturing the schedule of protein transport of membrane system in plant cell. Seven key tasks are emphasized in this paper in terms of the review on the effects and mechanisms of key ecological factors including water stress on plant nitrogen metabolism. They are: 1) the absorption mechanism of plant based on different nitrogen sources and environmental regulations, 2) the localization and compartmentalization of the key enzymes of nitrogen mechanism in plant cell, 3) the gene and environmental regulating model and their relationships in various key enzymes of nitrogen metabolism, 4) the function of main cell organs and their responses to environmental factors in nitrogen metabolism process, 5) physiological and chemical mechanism of nitrogen and the relationship between the mechanism and protein formation during crop grain filling, 6) improving gene structure of special species or cultivars using gene engineering methods to enhance the resistance to environmental factor stress and the efficiency of absorption and transportation of nitrogen, and 7) the mechanism of natural nitrogen cycle and its response to human activity disturbance.
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PMID:[Research advance in nitrogen metabolism of plant and its environmental regulation]. 1522 8

The Central Asian Taklamakan desert is characterized by a hyperarid climate with less than 50 mm annual precipitation but a permanent shallow groundwater table. The perched groundwater (2-16 m) could present a reliable and constant source of nitrogen throughout the growing season and help overcome temporal nitrogen limitations that are common in arid environments. We investigated the importance of groundwater and nitrogen fixation in the nitrogen metabolism of desert plants by assessing the possible forms and availability of soil N and atmospheric N and the seasonal variation in concentration as well as isotopic composition of plant N. Water availability was experimentally modified in the desert foreland through simulated flooding to estimate the contribution of surface water and temporally increased soil moisture for nutrient uptake and plant-water relations. The natural vegetation of the Taklamakan desert is dominated by plants with high foliar nitrogen concentrations (2-3% DM) and leaf nitrate reductase activity (NRA) (0.2-1 micromol NO2- g(-1) FW h(-1)). There is little evidence that nitrogen is a limiting resource as all perennial plants exhibited fast rates of growth. The extremely dry soil conditions preclude all but minor contributions of soil N to total plant N so that groundwater is suggested as the dominant source of N with concentrations of 100 microM NO3-. Flood irrigation had little beneficial effect on nitrogen metabolism and growth, further confirming the dependence on groundwater. Nitrogen fixation was determined by the 15N natural abundance method and was a significant component of the N-requirement of the legume Alhagi, the average contribution of biologically fixed nitrogen in Alhagi was 54.8%. But nitrogen fixing plants had little ecological advantage owing to the more or less constant supply of N available from groundwater. From our data we conclude that the perennial species investigated have adapted to the environmental conditions through development of root systems that access groundwater to satisfy demands for both water and nutrients. This is an ecologically favourable strategy since only groundwater is a predictable and stable resource.
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PMID:Nitrogen fixation and metabolism by groundwater-dependent perennial plants in a hyperarid desert. 1537 87

Nitrogen assimilation by plant symbiotic fungi plays a central role in the mutualistic interaction established by these organisms, as well as in nitrogen flux in a variety of soils. In the present study, we report on the functional properties, structural organization and distinctive mode of regulation of TbNrt2 (Tuber borchii NRT2 family transporter), the nitrate transporter of the mycorrhizal ascomycete T. borchii. As revealed by experiments conducted in a nitrate-uptake-defective mutant of the yeast Hansenula polymorpha, TbNrt2 is a high-affinity transporter (K(m)=4.7 microM nitrate) that is bispecific for nitrate and nitrite. It is expressed in free-living mycelia and in mycorrhizae, where it preferentially accumulates in the plasma membrane of root-contacting hyphae. The TbNrt2 mRNA, which is transcribed from a single-copy gene clustered with the nitrate reductase gene in the T. borchii genome, was specifically up-regulated following transfer of mycelia to nitrate- (or nitrite)-containing medium. However, at variance with the strict nitrate-dependent induction commonly observed in other organisms, TbNrt2 was also up-regulated (at both the mRNA and the protein level) following transfer to a nitrogen-free medium. This unusual mode of regulation differs from that of the adjacent nitrate reductase gene, which was expressed at basal levels under nitrogen deprivation conditions and required nitrate for induction. The functional and expression properties, described in the present study, delineate TbNrt2 as a versatile transporter that may be especially suited to cope with the fluctuating (and often low) mineral nitrogen concentrations found in most natural, especially forest, soils.
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PMID:Functional properties and differential mode of regulation of the nitrate transporter from a plant symbiotic ascomycete. 1620 72

Ammonia assimilation by the plastidic glutamine synthetase/glutamate synthase system requires 2-oxoglutarate (2-OG) as a carbon precursor. Plastids depend on 2-OG import from the cytosol. A plastidic dicarboxylate translocator 1-[2-OG/malate translocator (DiT1)] has been identified and its substrate specificity and kinetic constants have been analyzed in vitro. However, the role of DiT1 in intact plants and its significance for ammonia assimilation remained uncertain. Here, to study the role of DiT1 in intact plants, its expression was antisense-repressed in transgenic tobacco plants. This resulted in a reduced transport capacity for 2-OG across the plastid envelope membrane. In consequence, allocation of carbon precursors to amino acid synthesis was impaired, organic acids accumulated and protein content, photosynthetic capacity and sugar pools in leaves were strongly decreased. The phenotype was consistent with a role of DIT1 in both, primary ammonia assimilation and the re-assimilation of ammonia resulting from the photorespiratory carbon cycle. Unexpectedly, the in situ rate of nitrate reduction was extremely low in alpha-DiT1 leaves, although nitrate reductase (NR) expression and activity remained high. We hypothesize that this discrepancy between extractable NR activity and in situ nitrate reduction is due to substrate limitation of NR. These findings and the severe phenotype of the antisense plants point to a crucial role of DiT1 at the interface between carbon and nitrogen metabolism.
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PMID:Antisense repression reveals a crucial role of the plastidic 2-oxoglutarate/malate translocator DiT1 at the interface between carbon and nitrogen metabolism. 1636 65

Nitrogen is a vital component in living organisms as it participates in the making of essential biomolecules such as proteins, nucleic acids, etc. In the biosphere, nitrogen cycles between the oxidation states +V and -III producing many species that constitute the biogeochemical cycle of nitrogen. All reductive branches of this cycle involve the conversion of nitrate to nitrite, which is catalyzed by the enzyme nitrate reductase. The characterization of nitrate reductases from prokaryotic organisms has allowed us to gain considerable information on the molecular basis of nitrate reduction. Prokaryotic nitrate reductases are mononuclear Mo-containing enzymes sub-grouped as respiratory nitrate reductases, periplasmic nitrate reductases and assimilatory nitrate reductases. We review here the biological and molecular properties of these three enzymes along with their gene organization and expression, which are necessary to understand the biological processes involved in nitrate reduction.
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PMID:Bacterial nitrate reductases: Molecular and biological aspects of nitrate reduction. 1641 15

Experiments were carried out to examine the effects of nitrogen source on nitrogen incorporation into cyanophycin during nitrogen limitation and repletion, both with or without inhibition of protein synthesis, in cyanobacteria grown on either nitrate or ammonium. The use of nitrate and ammonium, 14N labeled in the growth medium and 15N labeled in the repletion medium, allows the determination of the source of nitrogen in cyanophycin using proton nuclear magnetic resonance spectroscopy. The data suggest that nitrogen from both the breakdown of cellular protein (14N) and directly from the medium (15N) is incorporated into cyanophycin. Nitrogen is incorporated into cyanophycin at different rates and to different extents, depending on the source of nitrogen (ammonium or nitrate) and whether the cells are first starved for nitrogen. These differences appear to be related to the activity of nitrate reductase in cells and to the possible expression of cyanophycin synthetase during nitrogen starvation.
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PMID:Effect of nitrogen source on cyanophycin synthesis in Synechocystis sp. strain PCC 6308. 1642 97

Nontarget injury from glyphosate drift is a concern among growers using non-glyphosate-resistant (non-GR) cultivars. The effects of glyphosate drift on nitrate assimilation and nitrogen fixation potential, nodule mass, and yield of non-GR soybean were assessed in a field trial at Stoneville, MS. A non-GR soybean cultivar 'Delta Pine 4748S' was treated with glyphosate at 12.5% of use rate of 0.84 kg of active ingredient/ha at 3 (V2), 6 (V7), and 8 (R2, full bloom) weeks after planting (WAP) soybean to simulate glyphosate drift. Untreated soybean was used as a control. Soybeans were sampled weekly for 2 weeks after each glyphosate treatment to assess nitrate assimilation and N2 fixation potential. Nitrate assimilation was assessed using in vivo nitrate reductase assay in leaves, stems, roots, and nodules. Nitrogen fixation potential was assessed by measuring nitrogenase activity using the acetylene reduction assay (ARA). Nitrogen content of leaves, shoots, and seed and soybean yield were also determined. In the first sampling date (4 WAP), glyphosate drift caused a significant decrease in NRA in leaves (60%), stems (77%), and nodules (50%), with no decrease in roots. At later growth stages, NRA in leaves was more sensitive to glyphosate drift than stems and roots. Nitrogenase activity was reduced 36-58% by glyphosate treatment at 3 or 6 WAP. However, glyphosate treatment at 8 WAP had no effect on nitrogenase activity. Nitrogen content was affected by glyphosate application only in shoots after the first application. No yield, seed nitrogen, protein, or oil concentration differences were detected. These results suggest that nitrate assimilation and nitrogen fixation potential were significantly reduced by glyphosate drift, with the greatest sensitivity early in vegetative growth. Soybean has the ability to recover from the physiological stress caused by glyphosate drift.
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PMID:Simulated glyphosate drift influences nitrate assimilation and nitrogen fixation in non-glyphosate-resistant soybean. 1663 96

Anabaena cylindrica grown with nitrate required higher levels of sodium (0.4 meq/l NaCl) to prevent chlorosis than when grown without combined nitrogen (0.004 meq/l NaCl). Nitrite accumulated in sodium-deficient cultures containing nitrate. Amounts of nitrite similar to those found in deficient cultures when added to normal cultures resulted in a chlorosis of the cells. Thus loss of chlorophyll was caused by nitrite toxicity.A deficiency of sodium resulted in an increased incorporation of (15)NO(3), (15)NO(2), (15)NH(3) or (14)C glutamate into protein compared with normal cells. The enzyme nitrate reductase was markedly increased in cells grown without sodium.Evidence from chloramphenicol treatment of the cells suggests that sodium may exert its control of nitrate reductase through a protein factor(s).By contrast, N(2) fixation was reduced in sodium deficient cells. Since the incorporation of ammonia or glutamate into protein was increased under these conditions, it is likely that the element is required for the conversion of N(2) gas into ammonia. Various nitrogenous compounds including ammonium chloride, amides and amino acids at low concentrations (0.1 mm) greatly reduced the nitrite accumulation in sodium-deficient cultures.
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PMID:Some Effects of Sodium on Nitrate Assimilation and N(2) Fixation in Anabaena cylindrica. 1665 97

Nitrate reductase activity and protein percentage of various tree parts of two Populus clones were determined in relation to nitrate ion activity. Nitrogen was supplied as NH(4)NO(3) in a nutriculture system. Wisconsin-5 had significantly greater nitrate reductase activity than Tristis No. 1. Protein percentages of leaf plastochron index 10 leaves (tenth leaf below first leaf lamina exceeding 20 mm in length), bottom leaves, and roots in relation to nitrate ion activity were not appreciably different between clones. The nitrate reductase activity and protein percentage of Tristis No. 1 apex started to level off at the same nitrate ion activity, about 0.09 mm. In Wisconsin-5 apex protein percentage continued to increase at nitrate ion activities where nitrate reductase activity decreases sharply, suggesting that protein nitrogen was being supplied by ammonium ion. The difference in nitrate reductase activity between clones was probably due to genetically determined ability to synthesize nitrate reductase in response to nitrate ion. The expression of nitrate reductase activity was not an index of nitrogen assimilation ability but may be a useful index of growth potential when nitrate ion does not limit nitrate reductase synthesis.
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PMID:Nitrate reductase activity and protein concentration of two populas clones. 1665 55


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