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)

Ralstonia eutropha (formerly Alcaligenes eutrophus) TF93 is pleiotropically affected in the translocation of redox enzymes synthesized with an N-terminal signal peptide bearing a twin arginine (S/T-R-R-X-F-L-K) motif. Immunoblot analyses showed that the catalytic subunits of the membrane-bound [NiFe] hydrogenase (MBH) and the molybdenum cofactor-binding periplasmic nitrate reductase (Nap) are mislocalized to the cytoplasm and to the inner membrane, respectively. Moreover, physiological studies showed that the copper-containing nitrous oxide reductase (NosZ) was also not translocated to the periplasm in strain TF93. The cellular localization of enzymes exported by the general secretion system was unaffected. The translocation-arrested MBH and Nap proteins were enzymatically active, suggesting that twin-arginine signal peptide-dependent redox enzymes may have their cofactors inserted prior to transmembrane export. The periplasmic destination of MBH, Nap, and NosZ was restored by heterologous expression of Azotobacter chroococcum tatA mobilized into TF93. tatA encodes a bacterial Hcf106-like protein, a component of a novel protein transport system that has been characterized in thylakoids and shown to translocate folded proteins across the membrane.
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PMID:Ralstonia eutropha TF93 is blocked in tat-mediated protein export. 1063 89

The NAD(P)H nitrate reductase (NR) from Chlamydomonas reinhardtii is encoded by the structural gene Nia1. Numerous data from the literature indicate that this enzyme is submitted to complex regulation mechanisms involving multiple controls at transcriptional and post-transcriptional levels. To specifically investigate the regulation of the Nia1 gene at the transcriptional level, NR+ and NR- transformed cells harbouring the Nia1:Ars construct (Nia1 promoter fused to the arylsulfatase (ARS)-encoding Ars reporter gene) were cultivated under various experimental conditions and the ARS activities were recorded. ARS levels were very low in cells grown in the presence of NH4Cl and dramatically increased on agar medium deprived of any nitrogen source or containing nitrate, nitrite, urea, arginine or glutamine. Compared to nitrogen-free medium, a slight positive effect of nitrate in the NR+ strain and a significant negative effect of nitrite in both NR+ and NR- strains were observed. The ARS activities were high in the light and very low in the dark or in the light in the presence of DCMU, indicating that Nia1 transcription is strikingly dependent on photosynthetic activity. Acetate used as a carbon source in the dark did not substitute for light in stimulating Nia1:Ars expression. Inactivation of NR by tungstate treatment of the NR+ strain resulted in a dramatic increase of ARS level suggesting that in Chlamydomonas, like in higher plants, active NR negatively regulates the transcription of the NR structural gene. Deleting the major part of the Nia1 leader sequence still present in the chimeric gene resulted in a decrease of ARS level but did not modify the regulation pattern.
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PMID:Transcriptional regulation of the Nia1 gene encoding nitrate reductase in Chlamydomonas reinhardtii: effects of various environmental factors on the expression of a reporter gene under the control of the Nia1 promoter. 1064 29

The periplasmic selenate reductase (Ser) of Thauera selennatis is a component of the electron transport chain catalyzing selenate reduction with acetate as the electron donor (i.e., selenate respiration). The purified enzyme consists of three subunits (SerA, SerB and SerC). Using transposon (i.e., Tn5) mutagenesis selenate reductase mutants were isolated. Junction fragments of DNA adjacent to the integrated Tn5 were used, together with oligonucleotides derived from the N-termini of SerA and SerB, to clone from a gene bank a DNA fragment that contained the corresponding genes. After sequencing, serA, serB and serC were identified by sequence comparison with the N-termini of the three subunits. The genes are arranged in the order serA, serB, serC; a fourth open reading frame (serD) in between, but overlapping serB and serC, is also present. The serA gene product contains an apparent leader peptide with a twin-arginine motif. The remainder of the translated amino acid sequence is similar to that of a number of prokaryotic molybdenum-containing enzymes (e.g., nitrate reductases and formate dehydrogenases of Escherichia coli). The serB gene product contains four cysteine clusters and is similar to various iron-sulfur protein subunits. The serC gene product contains a putative Sec-dependent leader peptide, but there are no similarities between the remainder of the translated protein and other protein subunits. The SerC contains two histidine and four methionine residues, and these may noncovalently bind heme b--which is a component of the active selenate reductase. The serD gene product encodes a putative protein that shows no significant sequence similarities to other proteins. However, the location of the serD within the other ser genes is similar to that of narJ within the E. coli narGHJI operon (nitrate reductase A); thus suggesting that the role of SerD may be similar to that of NarJ, which is a system-specific chaperone protein.
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PMID:Cloning and sequencing of the genes encoding the periplasmic-cytochrome B-containing selenate reductase of Thauera selenatis. 1082 93

Tobacco (Nicotiana tabacum L.) plants were subjected to a prolonged period of sulfur-deprivation to characterize molecular and metabolic mechanisms that permit control of primary N-metabolism under these conditions. Prior to the appearance of chlorotic lesions, sulfur-deprived tobacco leaves showed a strong decrease in the sulfate content and changes in foliar enzyme activities, mRNA accumulation and amino-acid pools. The basic amino acids glutamine, asparagine and arginine accumulated in the leaves of sulfur-deprived plants, while the foliar concentrations of aspartate, glutamate, serine or alanine remained fairly unchanged. Maximal extractable nitrate reductase (NR; EC 1.6.6.1) activity decreased strongly in response to sulfur-deprivation. The decrease in maximal extractable NR activity was accompanied by a decline in NR transcripts while the mRNAs of the plastidic glutamine synthetase (EC 6.1.3.2) or the beta-subunit of the mitochondrial ATP synthase were much less affected. Nitrate first accumulated in leaves of tobacco during sulfur-deprivation but then declined. An appreciable amount of nitrate was, however, present in severely sulfur-depleted leaves. The repression of NR gene expression is, therefore, not related to the decrease in the leaf nitrate level. However, glutamine- and/or asparagine-mediated repression of NR gene transcription is a possible mechanism of control in situations when glutamine and asparagine accumulate in leaves and provides a feasible explanation for the reduction in NR activity during sulfur-deprivation. The removal of reduced nitrogen from primary metabolism by redirection and storage as arginine, asparagine or glutamine combined with the down-regulation of nitrate reduction via glutamine- and/or asparagine-mediated repression of NR gene transcription may contribute to maintaining a normal N/S balance during sulfur-deprivation and indicate that the co-ordination of N- and S-metabolism is retained under these conditions.
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PMID:Negative regulation of nitrate reductase gene expression by glutamine or asparagine accumulating in leaves of sulfur-deprived tobacco. 1103 May 59

Numerous methods are available for measurement of nitrate (NO(-)(3)). However, these assays can either be time consuming or require specialized equipment (e.g., nitrate reductase, chemiluminescent detector). We have developed a method for simultaneous evaluation of nitrate and nitrite concentrations in a microtiter plate format. The principle of this assay is reduction of nitrate by vanadium(III) combined with detection by the acidic Griess reaction. This assay is sensitive to 0.5 microM NO(-)(3) and is useful in a variety of fluids including cell culture media, serum, and plasma. S-Nitrosothiols and L-arginine derivatives were found to be potential interfering agents. However, these compounds are generally minor constituents of biological fluids relative to the concentration of nitrate/nitrite. This report introduces a new, convenient assay for the stable oxidation products of nitrogen oxide chemistry in biological samples.
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PMID:A rapid, simple spectrophotometric method for simultaneous detection of nitrate and nitrite. 1600 57

Hydroponically grown spinach plants were deprived of an external source of sulphate after an initial period when the S-supply was sufficient. The time-course of events following this treatment was monitored. The first responses were found in the uptake and translocation of NO(3)(-) and the uptake of SO(4)(2-). The former declined by approximately 50%, the effect being most significant at higher [NO(3)(-)](ext.) while the latter increased 6-fold over a 4 d period. Growth in the absence of external SO(4)(2-) resulted in exhaustion of internal SO(4)(2-) pools, the effect being seen first in roots, then in young leaves and, after a marked delay, in mature leaves. In young leaves, there were dramatic increases in the [NO(3)(-)] and the content of arginine in the first 2 d of S-deprivation. The concentration of glutamine, the most abundant amino acid in S-sufficient conditions, also more than doubled in S-deficient young leaves. The changes in arginine levels were also found in older leaves, but the change in glutamine level was not seen. Assays of nitrate reductase activity (NRA) and nitrate reductase (NR) mRNA from young leaves of S-replete and S-deprived plants revealed a divergence in activity and content only late in the experiments (between days 4 and 8) when results were expressed on a unit leaf basis. However, there were also time-dependent changes in the protein content that kept the specific activities (NRA:protein and RNA:protein) more or less unchanged. The results imply that the impact of S-deficiency on N-utilization are more sensitively monitored by simple measurements of the chemical composition of young leaves than by measurements of NRA or NR transcript abundance. They also suggest that protein synthesis in young leaves is strongly dependent on a continuous supply of SO(4)(2-) from outside the plant.
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PMID:Rapid disruption of nitrogen metabolism and nitrate transport in spinach plants deprived of sulphate. 1118 20

To quantitatively characterise the nitrate reductase activity in the human oral cavity, a new assay based on holding 20 ml of 10 mg nitrate-N/L solution in the mouth was developed. The mouth assay appeared to relate primarily to the oral cavity surface rather than to the saliva. Nitrite formation in the assay was 50-100 times higher compared to in vitro incubation. In the proposed assay, the nitrite formation linearly increased over a period of 3 min. The average nitrate reductase activity in the oral cavity of 20 subjects was 2.39+/-1.52 microg nitrite-N formed/person x min. The nitrate reductase activity measured for two subjects at different hours varied about 15% for the same subject. The average nitrate reductase activity measured in June for 10 subjects (3.43+/-1.75 microg-N/person x min) was significantly higher than that measured in November for 10 other subjects (1.54+/-0.46 microg-N/person x min). Therefore, the nitrate reductase activity in the oral cavity appears to be influenced by the seasonal conditions. Although the amounts of nitrite formed in the mouth assay increased with increasing levels of nitrate, the rate of nitrate to nitrite reduction decreased with increasing levels of nitrate. The nitrite formation was also affected by the pH, with an optimal pH about 8. The nitrite formation was not influenced by uptake in the mouth of glucose, L-ascorbic acid and L-arginine.
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PMID:Quantitative measurement of the nitrate reductase activity in the human oral cavity. 1129 86

Six strains of Lactobacillus fermentum and Lactobacillus plantarum were investigated for nitric oxide (NO) production. First, the potential presence of NO synthase was examined. None of the strains of L. fermentum and L. plantarum examined produced NO from L-arginine under aerobic conditions. Interestingly, all L. fermentum strains expressed strong L-arginine deiminase activity. All L. fermentum strains produced NO in MRS broth, but the NO was found to be chemically derived from nitrite, which was produced by L. fermentum from nitrate present in the medium. Indeed all L. fermentum strains express nitrate reductase under anaerobic conditions. Moreover, one strain, L. fermentum LF1, had nitrate reductase activity under aerobic conditions. It was also found that L. fermentum strains JCM1173 and LF1 possessed ammonifying nitrite reductase. The latter strain also had denitrifying nitrite reductase activity at neutral pH under both anaerobic and aerobic conditions. The LF1 strain is thus capable of biochemically converting nitrate to NO. NO and nitrite produced from nitrate by lactobacilli may constitute a potential antimicrobial mechanism. studied in a rat acute liver injury model (Adawi et al. 1997). The results indicate that Lactobacillus plantarum DSM 9842 may possess NOS (Adawi et al. 1997). However, NO production from L-arginine has not been investigated in pure cultures of L. plantarum. According to the results of a 15N enrichment experiment, traces of (NO2-+NO3-)-N (total oxidised nitrogen: TON), which seemed to be formed by the resting cells of Lactobacillus fermentum IFO3956, appeared to be derived from L-arginine (Morita et al. 1997). Therefore, it was suggested that L. fermentum may possess a NOS. However, NO produced from L-arginine was not directly measured and a NOS inhibitor test was not performed by Morita et al. (1997). It is known that L-arginine deiminase (ADI) in bacteria may convert L-arginine to NH4+ (Cunin et al. 1986), which may be further oxidised to TON via nitrification by bacteria. Therefore, 15N enrichment experiments could not definitely conclude that L. fermentum possess NOS to convert L-arginine directly to NO. In this study, six Lactobacillus strains belonging to L. plantarum and L. fermentum were measured for NO production in MRS broth. The metabolism of nitrate and L-arginine by the Lactobacillus cell suspensions was also studied. The possibility that NO and nitrite production by lactobacilli may be a potential probiotic trait is also discussed.
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PMID:Evaluation of nitric oxide production by lactobacilli. 1154 28

Recently, transgenic potato plants were created showing underexpression of the 20R isoform of the 14-3-3 protein. The transgenic plants grown in tissue culture showed a significant increase in nitrate reductase activity and a decrease in nitrate level. The transgenic line with the lowest 14-3-3 quantity was field-trialed (1997-2000) and analyzed. The reduction in the 14-3-3 protein level consistently resulted in a starch content increase and in an increase in the ratio of soluble sugars to starch in the tubers, although the latter was only barely visible. The determination of amino acid composition in the tubers showed a significant increase in methionine, proline, and arginine content and a slight but consistent increase in hydrophobic amino acid and lysine content in the cells of the transgenic potato plants. We also observed an increase in the crude protein content, from 19 to 22.1% of the control value in consecutive years. It is proposed that all of these changes might have resulted from the downregulation of nitrate reductase and sucrose phosphate synthase activities by 14-3-3, although other potential mechanisms cannot be excluded (e.g., an increase in enzyme protein level). 14-3-3-repressed transgenic plants showed a significant increase in calcium content in their tubers. It is thus proposed that a function of the isolated 14-3-3 isoform is in the control of amino acid synthesis and calcium metabolism. However, the mechanism of this control is as yet unknown.
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PMID:Repression of the 14-3-3 gene affects the amino acid and mineral composition of potato tubers. 1190 69

Biological activity of nitric oxide (NO) production was investigated in the unicellular green alga Chlamydomonas reinhardtii. An NO specific electrode detected a rapid increase in signal when nitrite (NO(2)(-)) was added into a suspension of C. reinhardtii intact cells in the dark. The addition of KCN or the NO quencher bovine hemoglobin completely abolished the signal, verifying that the nitrite-dependent increase in signal is due to enzymatic NO production. L-arginine, the substrate for NO synthase, did not induce detectable NO production and the NOS inhibitor N(omega)-nitro-L-arginine showed no inhibitory effect on the nitrite-dependent production of NO. Illuminating cells showed a significant suppressive effect on NO production. When the photosynthetic electron transport inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea was present in the suspension, C. reinhardtii cells produced NO after the addition of nitrite even under illumination. Kinetic and microscopic observations, using the intracellular fluorescent NO probe 4,5-diaminofluorescein-2 diacetate, both demonstrated that NO was produced within the cells in response to the addition of nitrite. The Chlamydomonas mutant cc-2929, which lacks nitrate reductase (NR) activity, did not display any of the responses observed in the wild-type cells. The results presented here provide direct in vivo evidence to confirm that NR is involved in the nitrite-dependent NO production in the green alga.
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PMID:Nitric oxide production mediated by nitrate reductase in the green alga Chlamydomonas reinhardtii: an alternative NO production pathway in photosynthetic organisms. 1191 83


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