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)

NapC is a tetra-haem member of a family of bacterial membrane-anchored multi-haem c -type cytochromes implicated in electron transfer between membrane quinols and periplasmic enzymes. The water-soluble tetra-haem fragment of Paracoccus pantotrophus NapC has been expressed as a periplasmic protein (NapC(sol)) in Paracoccus denitrificans, P. pantotrophus and Escherichia coli. Site-specific mutagenesis of NapC(sol), combined with spectroscopic studies, suggests that each haem iron centre has bis -histidinyl co-ordination. Four proximal ligands arise from each of four Cys-Xaa-Xaa-Cys-His haem-binding motifs; candidates for the four distal ligands are His(81), His(99), His(174) and His(194). NapC(H81A), NapC(H99A), NapC(H174A) and NapC(H194A) mutants (with alanine substituted for each of the four candidate residues) have all been purified from E. coli. In each case, one of the haems has become high-spin, as judged by the presence of a broad absorption band between 620 nm and 650 nm for the oxidized cytochrome; this feature is absent for wild-type protein and presumably arises because of the absence of the distal histidine ligand from one of the haems. NapC(H81A) and NapC(H174A) are less well expressed in E. coli than NapC(H99A) and NapC(H194A) and cannot be detected when expressed in P. denitrificans or P. pantotrophus. In vitro and in vivo complementation studies demonstrate that the soluble periplasmic NapC can mediate electron transfer from quinols to the periplasmic nitrate reductase. This capacity was retained in vitro with the NapC(H99A) and NapC(H194A) mutants but was lost in vivo. A model for the structural organization of NapC(sol) into two domains, each containing a di-haem pair, is proposed. In this model, each haem pair obtains one distal haem ligand from its own domain and a second from the other domain. The suggestion of two domains is supported by observations that the 24 kDa NapC(sol) cleaves to yield a 12 kDa haem-staining band. Determination of the cleavage site showed it was between two equally sized di-haem domains predicted from sequence analysis.
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PMID:Identification of two domains and distal histidine ligands to the four haems in the bacterial c-type cytochrome NapC; the prototype connector between quinol/quinone and periplasmic oxido-reductases. 1218 31

We have generated a chromosomal mutant of moeB (moeBA228T) that demonstrates limited molybdenum cofactor (molybdo-bis(molybdopterin guanine dinucleotide) (Mo-bisMGD)) availability in Escherichia coli and have characterized its effect on the maturation and physiological function of two well-characterized respiratory molybdoenzymes: the membrane-bound dimethylsulfoxide (DMSO) reductase (DmsABC) and the membrane-bound nitrate reductase A (NarGHI). In the moeBA228T mutant strain, E. coli F36, anaerobic respiratory growth is possible on nitrate but not on DMSO, indicating that cofactor insertion occurs into NarGHI but not into DmsABC. Fluorescence analyses of cofactor availability indicate little detectable cofactor in the moeBA228T mutant compared with the wild-type, suggesting that NarGHI is able to scavenge limiting cofactor, whereas DmsABC is not. MoeB functions to sulfurylate MoaD, and in the structure of the MoeB-MoaD complex, Ala-228 is located in the interface region between the two proteins. This suggests that the moeBA228T mutation disrupts the interaction between MoeB and MoaD. In the case of DmsABC, despite the absence of cofactor, the twin-arginine signal sequence of DmsA is cleaved in the moeBA228T mutant, indicating that maturation of the holoenzyme is not cofactor-insertion dependent.
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PMID:Differential effects of a molybdopterin synthase sulfurylase (moeB) mutation on Escherichia coli molybdoenzyme maturation. 1223 97

This article reviews the relationship between the energy status of plant cells under O(2) stress (e.g. waterlogging) and the maintenance of membrane intactness, using information largely derived from suspension cultures of anoxia-intolerant potato cells. Energy-related parameters measured were fermentation end-products (ethanol, lactate, alanine), respiratory rate, ATP, adenylate energy charge, nitrate reductase activity and biomass. ATP synthesis rates were calculated from the first four parameters. Reactive oxygen species were estimated from H(2)O(2) and superoxide levels, and the enzymatic detoxification potential from the activity levels of catalase and superoxide dismutase. Structure-related parameters were total fatty acids, free fatty acids (FFAs), lipid hydroperoxides, total phospholipids, N-acylphosphatidylethanolamine (NAPE) and cell viability. The following issues are addressed in this review: (1) what is the impact of anoxia on membrane lipids and how does this relate to energy status; (2) does O(2) per se play a role in these changes; (3) under which conditions and to what extent does lipid peroxidation occur upon re-aeration; and (4) can the effects of re-aeration be distinguished from those of anoxia? The emerging picture is a reappraisal of the relative contributions of anoxia and re-aeration. Two successive phases (pre-lytic and lytic) characterize potato cells under anoxia. They are connected by a threshold in ATP production rate, below which membrane lipids are hydrolysed to FFAs, and NAPE increases. Since lipid peroxidation occurs only when cells are reoxygenated during the lytic phase, its biological relevance in an already damaged system is questionable.
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PMID:Impact of oxygen stress and energy availability on membrane stability of plant cells. 1232 74

The Neurospora crassa genome database was searched for sequence similarity to crnA, a nitrate transporter in Aspergillus nidulans. A 3.9-kb fragment (contig 3.416, subsequence 183190-187090) was cloned by PCR. The gene coding for this nitrate transporter was termed nit-10. The nit-10 gene specifies a predicted polypeptide containing 541 amino acids with a molecular mass of 57 kDa. In contrast to crnA, which is clustered together with niaD, encoding nitrate reductase, and niiA, encoding nitrite reductase, nit-10 is not linked to nit-3 (nitrate reductase), nit-6 (nitrite reductase), or to nit-2, nit-4 (both are positive regulators of nit-3), or nmr (negative regulator of nit-3) in Neurospora crassa. A nit-10 rip mutant failed to grow in the medium when nitrate (< 10 mM) was used as the sole nitrogen source, but grew similarly to wild type when nitrate concentration was 10 mM or higher. In addition, it showed strong sensitivity to cesium in the presence of nitrate and resistance to chlorate in the presence of alanine, proline, or hypoxanthine. The expression of nit-10 required nitrate induction and was subject to repression by nitrogen metabolites such as glutamine. Expression of nit-10 also required functional products of nit-2 and nit-4. The half-life of nit-10 mRNA was determined to be approximately 2.5 min.
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PMID:Identification and characterization of a nitrate transporter gene in Neurospora crassa. 1506 36

Posttranslational regulation of nitrate assimilation was studied in the cyanobacterium Synechocystis sp. strain PCC 6803. The ABC-type nitrate and nitrite bispecific transporter encoded by the nrtABCD genes was completely inhibited by ammonium as in Synechococcus elongatus strain PCC 7942. Nitrate reductase was insensitive to ammonium, while it is inhibited in the Synechococcus strain. Nitrite reductase was also insensitive to ammonium. The inhibition of nitrate and nitrite transport required the PII protein (glnB gene product) and the C-terminal domain of NrtC, one of the two ATP-binding subunits of the transporter, as in the Synechococcus strain. Mutants expressing the PII derivatives in which Ala or Glu is substituted for the conserved Ser49, which has been shown to be the phosphorylation site in the Synechococcus strain, showed ammonium-promoted inhibition of nitrate uptake like that of the wild-type strain. The S49A and S49E substitutions in GlnB did not affect the regulation of the nitrate and nitrite transporter in Synechococcus either. These results indicated that the presence or absence of negative electric charge at the 49th position does not affect the activity of the PII protein to regulate the cyanobacterial ABC-type nitrate and nitrite transporter according to the cellular nitrogen status. This finding suggested that the permanent inhibition of nitrate assimilation by an S49A derivative of PII, as was previously reported for Synechococcus elongatus strain PCC 7942, is likely to have resulted from inhibition of nitrate reductase rather than the nitrate and nitrite transporter.
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PMID:Posttranslational regulation of nitrate assimilation in the cyanobacterium Synechocystis sp. strain PCC 6803. 1562 21

Isolated cotyledons of fenugreek (Trigonella foenum graecum L.), which respond rapidly and specifically to the application of cytokinins with stimulated expansion, have been used to study the primary action of kinetin. Gross chemical analysis showed that ribonucleic acid increased within 24 hours in response to kinetin application. 8-Azaguanine inhibited both kinetin-induced expansion and RNA synthesis; 5-fluorodeoxyuridine inhibited only the RNA synthesis.Cotyledons produced nitrate reductase activity in response to 20 mm nitrate only in the presence of either light or kinetin and especially in the presence of both. Abscisic acid and inhibitors of RNA and protein synthesis depressed this response. Inhibitors affecting chloroplast development and function did not reduce the response in the presence of light and kinetin.In vitro incorporation of (14)C-l-leucine and (14)C-l-phenyl-alanine into protein by various recombinations of microsomal and 160,000g supernatant fractions varied according to the pretreatment which the cotyledons had received before the preparation of the fractions. Stimulatory effects were mainly associated with the microsomal fractions.The formation of leucine-, valine-, and tyrosine-tRNA complexes by high speed supernatant fractions from differently pretreated cotyledons was also compared. The sharp stimulation of the process by adding tRNA was found to be independent of the kind of preincubation that the cotyledons used for the tRNA extraction had received.It is concluded that the evidence is not in favor of kinetin correcting specific tRNA deficiencies. Kinetin removes a limitation that prevents the synthesis of RNA and genome expression.
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PMID:Action of kinetin on cotyledons of fenugreek. 1665 78

A nitrate uptake system is induced (along with nitrate reductase) when NH(4) (+)-grown Penicillium chrysogenum is incubated with inorganic nitrate in synthetic medium in the absence of NH(4) (+). Nitrate uptake and nitrate reduction are probably in steady state in fully induced mycelium, but the ratios of the two activities are not constant during the induction period. Substrate concentrations of ammonium cause a rapid decay of nitrate uptake and nitrate reductase activity. The two activities are differentially inactivated (the uptake activity being more sensitive). Glutamine and asparagine are as effective as NH(4) (+) in suppressing nitrate uptake activity. Glutamate and alanine were about half as effective as NH(4) (+). Cycloheximide interferes with the NH(4) (+)-induced decay of nitrate uptake activity. The ammonium transport system is almost maximally deinhibited (or derepressed) in nitrate-grown mycelium.
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PMID:Regulation of Nitrate Uptake in Penicillium chrysogenum by Ammonium Ion. 1665 63

The induction of nitrate reductase activity in root tips of cotton (Gossypium hirsutum L.) was regulated by several amino acids and by ammonium. Glycine, glutamine, and asparagine strongly inhibited induction of activity by nitrate and also decreased growth of sterile-cultured roots on a nitrate medium. Methionine, serine, and alanine weakly inhibited induction, and 11 other amino acids had little or no effect. Ammonium also decreased induction in root tips, but was most effective only at pH 7 or higher. The optimum conditions for ammonium regulation of induction were identical to those for growth of sterile-cultured roots on ammonium as the sole nitrogen source. Aspartate and glutamate strongly stimulated induction, but several lines of evidence indicated that the mechanism of this response was different from that elicited by the other amino acids. The effects of amino acids on induction appeared to be independent of nitrate uptake.In green shoot tissues, all attempts to demonstrate regulation of induction by amino acids failed. The great difference in observed responses of root and shoot to amino acids suggests that their nitrate reductase activities are regulated differently. Differential regulation of this enzyme is consistent with the responses of root and shoot nitrate reductase activity to nitrate.
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PMID:Differential regulation of nitrate reductase induction in roots and shoots of cotton plants. 1665 46

Excised 7-day-old oat (Avena sativa L. cv. Jaycee) leaves were incubated in media containing 7.1 millimolar KNO(3) and 0.15 millimolar tabtoxin or 1 millimolar methionine sulfoximine (MSO) to investigate the sources of the observed ammonium accumulated. Tabtoxin and MSO are known inhibitors of glutamine synthetase, the first enzyme in the primary pathway of ammonium assimilation. During a 4- to 6-hour incubation, there was little net change in protein or total amino acid concentration. Alanine, aspartate/asparagine, and glutamate/glutamine decreased markedly under these treatments, whereas several other amino acids increased. Exogenous (15)N from K(15)NO(3) was taken up and incorporated into the nitrate and ammonium fractions of leaves treated with tabtoxin or MSO. This result and the high in vitro activities of nitrate reductase indicated that reduction of nitrate was one source of the accumulated ammonium. Leaves incubated under 2% O(2) to reduce photorespiration accumulated only about 13% as much ammonium as did those under normal atmospheres. We conclude that most of the tabtoxin- or MSO-induced ammonium came from photo-respiration, and the remainder was from nitrate reduction.
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PMID:Sources of ammonium in oat leaves treated with tabtoxin or methionine sulfoximine. 1666 6

The present study shows for the first time the influence of exogenously applied amino acids and cytokinin on the physiological and molecular aspects of N metabolism in poplar trees. In a short-term feeding experiment, glutamine or trans-zeatin riboside (tZR) was added directly to the nutrient solution. NO3- net uptake declined significantly in response to both treatments. Feeding with glutamine brought about an increase in concentrations of different amino compounds in the roots (glutamine, glutamate, alanine, gamma-amino butyric acid (GABA) and NH4+, which negatively correlated with the net NO3- uptake. The plants showed a reduction of cytosolic glutamine synthetase 1 (GS1) transcript level in the roots. In addition, glutamine feeding changed the root-to-shoot distribution on N assimilation in favour of the leaves and plant internal N cycling. tZR treatment resulted in expansion of zeatin-type (Z-type) cytokinins in the roots and increased nitrate reductase (NR)-mRNA level. The results indicate that both particular amino acids and active cytokinins are involved in the feedback regulation of N uptake and metabolism in poplar. We propose that inhibition of N uptake by cytokinins in poplar is more complex than that mediated by amino compounds, and other effectors are involved in this regulation.
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PMID:Exogenous supply of glutamine and active cytokinin to the roots reduces NO3- uptake rates in poplar. 1708 Sep 50


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