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:1.7.1.2 (nitrate reductase)
3,861 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A cDNA, hvst1, was isolated from Hordeum vulgare by heterologous complementation in Escherichia coli. This cDNA encodes a high-affinity sulfate transporter that is 2442 bp in length and consists of 660 amino acids. Under steady-state conditions of sulfate supply during culture, sulfate influx (measured at 100 microM external sulfate concentration) and hvst1 transcript level were inversely correlated with sulfate concentrations in the culture solution. Glutathione (GSH) concentrations increased as external sulfate was increased from 2.5 to 250 microM. A time-course study, designed to investigate effects of sulfate withdrawal on the abundance of hvst1 transcript, showed a 5-fold increase of the latter within the first two hours. This was followed by a further slight increase during the next 46 h. These changes were accompanied by a parallel increase in sulfate influx and a decrease of root GSH concentrations. When plants that had been deprived of sulfate for 24 h were exposed to L-cysteine (Cys) or GSH for 3 h, GSH was the more effective down-regulator, reducing hvst1 transcript level to below that of unstarved controls. The decrease in transcript abundance induced by sulfate or Cys was partially relieved by the addition of buthionine sulfoximine (BSO), an inhibitor of GSH synthesis. Both hvst1 transcripts and sulfate influx increased as a function of N supply to N-starved plants. Amino oxyacetate acid (AOA), an aminotransferase inhibitor, when supplied with NO3-, increased transcript abundance of hvst1, while tungstate, methionine sulfoximine (MSO) and azaserine (AZA), inhibitors of nitrate reductase, glutamine synthetase and glutamate synthase (GOGAT), respectively, were without effect. AOA decreased root concentrations of aspartate (Asp), Cys and GSH; in contrast, glutamate (Glu) concentrations remained unchanged.
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PMID:Regulation of the hvst1 gene encoding a high-affinity sulfate transporter from Hordeum vulgare. 1048 22

A method is presented to isolate mutants of Chlorella sorokiniana with defects in NO3- metabolism. Three nitrite-reductase (NIR; E.C.1.7.7.1)-deficient mutants were obtained from 500 pinpoint-colony-forming clones. The final screening was performed using NO3-, NO2- or NH4+ as N-source. The mutants isolated absorb NO3- with rates close to those measured for the wild type and they excrete NO2- into the medium. The ratio between NO3- uptake and NO2- excretion was 1:1. The sensitivity of NO3- uptake to NH4+ was reduced in the mutant strains as it was in the N-starved wild type of Chlorella. Nitrate reductase (NR; EC 1.6.6.1) expression and NR activity were slightly reduced compared to the wild type due to feedback regulation in the mutant strains. No NIR protein was found in the three mutants. However, NIR activity was obtained (50% of the wild-type) for one mutant strain. The NIR-deficient mutants and the already available NR-deficient mutants will be promising tools for investigations of the nitrate assimilation pathway on the molecular level and for studies searching for signaling of C and N metabolism by inorganic N-compounds.
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PMID:Isolation and characterization of nitrite-reductase-deficient mutants of Chlorella sorokiniana (strain 211-8k). 1098 64

We examined changes in root system architecture and physiology and whole-plant patterns of nitrate reductase (NR) activity in response to atmospheric CO2 enrichment and N source to determine how changes in the form of N supplied to plants interact with rising CO2 concentration ([CO2]). Seedlings of Betula alleghaniensis Britt. and Pinus strobus L., which differ in growth rate, root architecture, and the partitioning of NR activity between leaves (Betula) and roots (Pinus), were grown in ambient (400 microl l(-1)) and elevated (800 microl l(-1)) [CO2] and supplied with either nitrate (NO3-) or ammonium (NH4+) as their sole N source. After 15 weeks of growth, plants were harvested and root system architecture, N uptake kinetics, and NR activity measured. Betula alleghaniensis responded to elevated [CO2] with significant increases in growth, regardless of the source of N. Pinus strobus showed no significant response in biomass production or allocation to elevated [CO2]. Both species exhibited significantly greater growth with NH4+ than with NO3-, along with lower root:shoot biomass ratios. Betula showed significant increases in total root length in response to elevated [CO2]. However, root N uptake rates in Betula (for both NO3- and NH4+) were either reduced or unchanged by elevated [CO2]. Pinus showed the opposite response to elevated [CO2], with no change in root architecture, but an increase in maximal uptake rates in response to elevated [CO2]. Nitrate reductase activity (on a mass basis) was reduced in leaves of Betula in elevated [CO2], but did not change in other tissues. Nitrate reductase activity was unaffected by elevated [CO2] in Pinus. Scaling this response to the whole-plant, NR activity was reduced in elevated [CO2] in Betula but not in Pinus. However, because Betula plants were larger in elevated [CO2], total whole-plant NR activity was unaffected.
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PMID:Ammonium and nitrate acquisition by plants in response to elevated CO2 concentration: the roles of root physiology and architecture. 1130 44

An understanding of root system capacity to acquire nitrogen (N) is critical in assessing the long-term growth impact of rising atmospheric CO2 concentration ([CO2]) on trees and forest ecosystems. We examined the effects of mycorrhizal inoculation and elevated [CO2] on root ammonium (NH4+) and nitrate (NO3-) uptake capacity in sweetgum (Liquidambar styraciflua L.) and loblolly pine (Pinus taeda L.). Mycorrhizal treatments included inoculation of seedlings with the arbuscular mycorrhizal (AM) fungus Glomus intraradices Schenck & Smith in sweetgum and the ectomycorrhizal (EM) fungus Laccaria bicolor (Maire) Orton in loblolly pine. These plants were then equally divided between ambient and elevated [CO2] treatments. After 6 months of treatment, root systems of both species exhibited a greater uptake capacity for NH4+ than for NO3-. In both species, mycorrhizal inoculation significantly increased uptake capacity for NO3-, but not for NH4+. In sweetgum, the mycorrhizal effect on NO3- and NH4+ uptake capacity depended on growth [C02]. Similarly, in loblolly pine, the mycorrhizal effect on NO3- uptake capacity depended on growth [CO2], but the effect on NH4+ uptake capacity did not. Mycorrhizal inoculation significantly enhanced root nitrate reductase activity (NRA) in both species, but elevated [CO2] increased root NRA only in sweetgum. Leaf NRA in sweetgum did not change significantly with mycorrhizal inoculation, but increased in response to [CO2]. Leaf NRA in loblolly pine was unaffected by either treatment. The results indicate that the mycorrhizal effect on specific root N uptake in these species depends on both the form of inorganic N and the mycorrhizal type. However, our data show that in addressing N status of plants under high [CO2], reliable prediction is possible only when information about other root system adjustments (e.g., biomass allocation to fine roots) is simultaneously considered.
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PMID:Influence of elevated CO2 and mycorrhizae on nitrogen acquisition: contrasting responses in Pinus taeda and Liquidambar styraciflua. 1130 52

Regulation of root N uptake by whole-plant signalling of N status was investigated at the molecular level in Arabidopsis thaliana plants through expression analysis of AtNrt2.1 and AtAmt1.1. These two genes encode starvation-induced high-affinity NO3- and NH4+ transporters, respectively. Split-root experiments indicate that AtNrt2.1 expression is controlled by shoot-to-root signals of N demand. Together with 15NO3- influx, the steady-state transcript level of this gene is increased in NO3--fed roots in response to N deprivation of another portion of the root system. Thus AtNrt2.1 is the first identified molecular target of the long-distance signalling informing the roots of the whole plant's N status. In contrast, AtAmt1.1 expression is predominantly dependent on the local N status of the roots, as it is mostly stimulated in the portion of the root system directly experiencing N starvation. The same behaviour was found for NH4+ influx, suggesting that the NH4+ uptake system is much less efficient than the NO3- uptake system, to compensate for a spatial restriction of N availability. Other major differences were found between the regulations of AtNrt2.1 and AtAmt1.1 expression. AtNrt2.1 is strongly upregulated by moderate level of N limitation, while AtAmt1.1 transcript level is markedly increased only under severe N deficiency. Unlike AtNrt2.1, AtAmt1.1 expression is not stimulated in a nitrate reductase-deficient mutant after transfer to NO3- as sole N source, indicating that NO3- per se acts as a signal repressing transcription of AtAmt1.1. These results reveal two fundamentally different types of mechanism involved in the feedback regulation of root N acquisition by the N status of the plant.
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PMID:Differential regulation of the NO3- and NH4+ transporter genes AtNrt2.1 and AtAmt1.1 in Arabidopsis: relation with long-distance and local controls by N status of the plant. 1138 56

An integrated enzyme-functionalized field-effect transistor (ENFET) device for the sensing of nitrate ions is described. An aminosiloxane-functionalized gate interface is modified with N-methyl-N'-(carboxyalkyl)-4,4'-bipyridinium relay units. The complex formed between nitrate reductase and the bipyridinium units on the gate surface is crosslinked with glutaric dialdehyde to yield a stable relay-enzyme layer on the gate interface. In the presence of sodium dithionite as electron donor, the biocatalyzed reduction of nitrate to nitrite ion is stimulated. The ratio between the oxidized and reduced states of the bipyridinium sites regulates the gate potential, and is controlled by the concentration of NO3- ions in the system. The effect of the chain length tethering the N-methyl-N'-(carboxyalkyl)-4,4'-bipyridinium units to the gate surface on the biocatalyzed reduction of NO3- ions, and on the NO3- FET sensor performance is discussed. The devices that include the bipyridinium units tethered to the gate interface with methylene chain length, -(CH2)n, where n > or = 7, reveal a detection limit of 7 x 10(-5) M for nitrate and a sensitivity of 52 +/- 2 mV dec-1. The response time of the device is as low as 50 s, and the operational time of the system is ca. 85 s. We estimate the surface coverage of nitrate reductase on the gate surface to be ca. 1.2 x 10(-12) mol cm-2.
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PMID:An integrated relay/nitrate reductase field-effect transistor for the sensing of nitrate (NO3-). 1139 8

Transformed tobacco (Nicotiana tabacum L.) plants with varying activities of the key enzyme of ammonia assimilation, ferredoxin-glutamine-alpha-ketoglutarate aminotransferase (Fd-GOGAT; EC 1.4.7.1), were used to examine the roles of ammonium, glutamine (Gln) and alpha-ketoglutarate (alpha-KG) in the regulation of nitrate reductase (NR; EC 1.6.6.1) transcript abundance. In wild-type leaf discs, NR mRNA abundance was increased following feeding with NO3-, sucrose and alpha-KG and decreased by feeding Gln. In air, leaves with decreased GOGAT accumulated Gln and alpha-KG simultaneously; this was accompanied by increased NR transcripts. The inhibition of NR transcription by Gln observed in leaf-disc experiments was therefore not observed in the low-Fd-GOGAT plants that accumulate Gln in vivo. The results suggest that the negative effect of Gln on NR transcript abundance was offset by high alpha-KG and that the relative amounts of alpha-KG and Gln are more important in controlling NR gene transcription than the concentration of either metabolite alone.
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PMID:Glutamine and alpha-ketoglutarate are metabolite signals involved in nitrate reductase gene transcription in untransformed and transformed tobacco plants deficient in ferredoxin-glutamine-alpha-ketoglutarate aminotransferase. 1146 92

Membrane associated nitrate reductase (NR) was detected in plasma membrane (PM) fractions isolated by aqueous two-phase partitioning from barley (Hordeum vulgare L. var CM 72) roots. The PM associated NR was not removed by washing vesicles with 500 millimolar NaCl and 1 millimolar EDTA and represented up to 4% of the total root NR activity. PM associated NR was stimulated up to 20-fold by Triton X-100 whereas soluble NR was only increased 1.7-fold. The latency was a function of the solubilization of NR from the membrane. NR, solubilized from the PM fraction by Triton X-100 was inactivated by antiserum to Chlorella sorokiniana NR. Anti-NR immunoglobulin G fragments purified from the anti-NR serum inhibited NO3- uptake by more than 90% but had no effect on NO2- uptake. The inhibitory effect was only partially reversible; uptake recovered to 50% of the control after thorough rinsing of roots. Preimmune serum immunoglobulin G fragments inhibited NO3- uptake 36% but the effect was completely reversible by rinsing. Intact NR antiserum had no effect on NO3- uptake. The results present the possibility that NO3- uptake and NO3- reduction in the PM of barley roots may be related.
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PMID:Inhibition of nitrate transport by anti-nitrate reductase IgG fragments and the identification of plasma membrane associated nitrate reductase in roots of barley seedlings. 1153 34

The effect of SeO3= and SeO4= on NO3- assimilation in 8-d-old barley (Hordeum vulgare L.) seedlings was studied over a 24-h period. Selenite at 0.1 mol m-3 in the uptake solutions severely inhibited the induction of NO3- uptake and active nitrate reductases. Selenate, at 1.0 mol m-3 in the nutrient solution, had little effect on induction of activities of these systems until after 12 h; however, when the seedlings were pretreated with 1.0 mol m-3 SeO4= for 24 h, subsequent NO3- uptake from SeO4(=) -free solutions was inhibited about 60%. Sulphate partially alleviated the inhibitory effect of SeO3= when supplied together in the ambient solutions, but had no effect in seedlings pretreated with SeO3=. By contrast, SO4= partially alleviated the inhibitory effect of SeO4= even in seedlings pretreated with SeO4=. Since uptake of NO3- by intact seedlings was also inhibited by SO3=, the percentage of the absorbed NO3- that was reduced was not affected. By contrast, SeO4=, which affected NO3- uptake much less, inhibited the percentage reduced of that absorbed. However, when supplied to detached leaves, both SeO3= and SeO4= inhibited the in vivo reduction of NO3- as well as induction of nitrate reductase and nitrite reductase activities. Selenite was more inhibitory than SeO4= ; approximately a five to 10 times higher concentration of SeO4= than SeO3= was required to achieve similar inhibition. In detached leaves, the inhibitory effect of both SeO3= and SeO4= on in vivo NO3- reduction as well as on the induction of nitrate reductase activity was partially alleviated by SO4=. The inhibitory effects of Se salts on the induction of the nitrite reductase were, however, completely alleviated by SO4=. The results show that in barley seedlings SeO3= is more toxic than SeO4=. The reduction of SeO4= to SeO3= may be a rate limiting step in causing Se toxicity.
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PMID:Comparative effects of selenite and selenate on nitrate assimilation in barley seedlings. 1153 99

The nature of the association between nitrate reductase (NR) and membranes was examined. Nitrate reductase activity (NRA) associated with the microsomal fraction of barley (Hordeum vulgare L.) roots amounted to 0.6 to 0.8% of soluble NRA following sonication in the presence of 250 mM KI and repeated osmotic shock. This treatment removed all contaminating soluble NRA from microsomes of uninduced barley roots that had been homogenized in a soluble extract from roots of NO3(-)-induced plants. On continuous sucrose gradients, NRA co-migrated specifically with VO4(-)-sensitive ATPase activity, a plasma membrane (PM) marker; activity of glucose-6-phosphate dehydrogenase, assayed as cytosolic marker, co-migrated with NRA. Microsomal NRA was absent in barley deficient in soluble NR. Perturbation and trypsinolysis experiments with PM vesicles isolated by aqueous two-phase partitioning indicated that NR is associated with the periphery of the cytoplasmic face of the bilayer. These results demonstrate that PM and soluble NRs are essentially the same protein but that the membrane-associated form is tightly bound. Although it is possible that PM-associated NR exists in vivo, unequivocal evidence for this has yet to be shown. However, PM NR is definitely present in vitro.
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PMID:Characterization of the association of nitrate reductase with barley (Hordeum vulgare L.) root membranes. 1153


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