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)

We have potentiometrically characterized the two hemes of Escherichia coli nitrate reductase A (NarGHI) using EPR and optical spectroscopy. NarGHI contains two hemes, a low-potential heme b(L) (E(m,7) = 20 mV; g(z)() = 3.36) and a high-potential heme b(H) (E(m, 7) = 120 mV; g(z)() = 3.76). Potentiometric analyses of the g(z)() features of the heme EPR spectra indicate that the E(m,7) values of both hemes are sensitive to the menaquinol analogue 2-n-heptyl-4-hydroxyquinoline N-oxide (HOQNO). This inhibitor causes a potential-inversion of the two hemes (for heme b(L), E(m,7) = 120 mV; for heme b(H), E(m,7) = 60 mV). This effect is corroborated by optical spectroscopy of a heme b(H)-deficient mutant (NarGHI(H56R)) in which the heme b(L) undergoes a DeltaE(m,7) of 70 mV in the presence of HOQNO. Another potent inhibitor of NarGHI, stigmatellin, elicits a moderate heme b(L) DeltaE(m,7) of 30 mV, but has no detectable effect on heme b(H). No effect is elicited by either inhibitor on the line shape or the E(m,7) values of the [3Fe-4S] cluster coordinated by NarH. When NarI is expressed in the absence of NarGH [NarI(DeltaGH)], two hemes are detected in potentiometric titrations with E(m,7) values of 37 mV (heme b(L); g(z)() = 3.15) and -178 mV (heme b(H); g(z)() = 2.92), suggesting that heme b(H) may be exposed to the aqueous milieu in the absence of NarGH. The identity of these hemes was confirmed by recording EPR spectra of NarI(DeltaGH)(H56R). HOQNO binding titrations followed by fluorescence spectroscopy suggest that in both NarGHI and NarI(DeltaGH), this inhibitor binds to a single high-affinity site with a K(d) of approximately 0.2 microM. These data support a functional model for NarGHI in which a single dissociable quinol binding site is associated with heme b(L) and is located toward the periplasmic side of NarI.
Biochemistry 1999 Sep 28
PMID:The hemes of Escherichia coli nitrate reductase A (NarGHI): potentiometric effects of inhibitor binding to narI. 1050 45

A single Hansenula anomala genomic DNA fragment containing the genes H.a.YNR1 (yeast nitrate reductase) and H.a.YNI1 (yeast nitrite reductase) encoding nitrate and nitrite reductase, respectively, was isolated from a lambda EMBL3 genomic DNA library. As probe, a 3.2 kb DNA fragment isolated from a lambda gt11 H. anomala genomic DNA library screened with antiserum anti-NR from H. anomala was used. H. a.YNR1 and H.a.YNI1 genes are separated by 473 bp and encode putative proteins of 870 and 1077 amino acids, respectively, with great similarity to nitrate and nitrite reductases from other organisms. Northern blot analysis revealed that both genes are highly expressed in nitrate, very low in nitrate plus ammonium, and no expression was detected in ammonium or nitrogen-free media. Levels of nitrate reductase and nitrite reductase were very low or undetectable by Western blot analysis in nitrogen-free and ammonium media, whereas both proteins were present in nitrate and ammonium plus nitrate media. The nucleotide sequence Accession No. is AF123281.
Yeast 2000 Sep 15
PMID:Cloning, sequencing, and expression of H.a.YNR1 and H.a.YNI1, encoding nitrate and nitrite reductases in the yeast Hansenula anomala. 1095 81

Mycobacterium avium subsp. paratuberculosis is a pathogen that causes chronic inflammation of the intestine in many animals, including primates, and is implicated in Crohn's disease in humans. It differs from other members of the M. avium complex in having 14-18 copies of IS900 inserted into conserved loci in its genome. In the present study, genomic DNA flanking 14 of these insertions was characterized and homologues in the Mycobacterium tuberculosis and M. avium subsp. avium genomes were identified. These included regions encoding a sigma factor (sigJ) at locus 3, a nitrate reductase (nirA) at locus 4, a transcription regulator (tetR) and polyketide synthase at locus 6, and a 6-O-methylguanine methyltransferase at locus 9. In addition, locus numbers were assigned to 9 of 15 RFLP bands previously described. IS900 insertion at 7 of the 14 characterized loci was into the RBS of a gene substituting an RBS encoded by IS900 sited two bases closer to the initiation codon. IS900 insertion at five loci interrupted an ORF at the target site, one of which encoded a homologue of the immunodominant mycobacterial DesA1 protein. Eleven of eighty-one M. avium subsp. paratuberculosis isolates lacked the insertion site at locus 6 together with flanking genomic DNA. This region was also absent from seven reference strains of M. avium subsp. avium, from one M. avium subsp. silvaticum and from six other mycobacterial species. A multiplex PCR of IS900 loci (MPIL) typing method was developed which was able to discriminate 10 different types of M. avium subsp. paratuberculosis from the panel of 81 isolates with consistent differences between those of bovine and ovine origin. Nine MPIL types corresponded with a single PstI/Bst:EII RFLP type, suggesting that this method may be applicable to typing of M. avium subsp. paratuberculosis directly from a sample without the need for culture. The remaining MPIL type corresponded with seven PstI/BstEII RFLP types. Further resolution of these may come from sequencing the remaining four uncharacterized IS900 loci.
Microbiology (Reading) 2000 Sep
PMID:Characterization of IS900 loci in Mycobacterium avium subsp. paratuberculosis and development of multiplex PCR typing. 1097 6

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.
Planta 2000 Sep
PMID:Negative regulation of nitrate reductase gene expression by glutamine or asparagine accumulating in leaves of sulfur-deprived tobacco. 1103 May 59

Expression of the dimethylsulfoxide respiratory (dor) operon of Rhodobacter is regulated by oxygen, light intensity and availability of substrate. Since dimethylsulfoxide reductase contains a pterin molybdenum cofactor, the role of molybdate in the regulation of dor operon expression was investigated. In this report we show that the molybdate-responsive transcriptional regulator, MopB, and molybdate are essential for maximal dimethylsulfoxide reductase activity and expression of a dorA::lacZ transcriptional fusion in Rhodobacter capsulatus. In contrast, mop genes are not required for the expression of the periplasmic nitrate reductase or xanthine dehydrogenase in R. capsulatus under conditions of molybdenum sufficiency. This is the first report demonstrating a clear functional difference between the ModE homologues MopB and MopA in this bacterium. The results suggest that MopA is primarily involved in the regulation of nitrogen fixation gene expression in response to molybdate while MopB has a role in nitrogen fixation and dimethylsulfoxide respiration.
FEMS Microbiol Lett 2000 Sep 15
PMID:Molybdate-dependent expression of dimethylsulfoxide reductase in Rhodobacter capsulatus. 1103 80

To assess the role of 14-3-3 proteins in the magnesium-dependent inhibition of nitrate reductase (NR) we tested the effect of magnesium on NR binding to 14-3-3s by coimmunoprecipitation and gel filtration. The stability of the 14-3-3 complex of NR was, unlike its activity, unaffected by magnesium. We therefore conclude that binding to 14-3-3s per se does not inhibit NR. Magnesium inhibited 14-3-3-bound NR much more strongly than 14-3-3-free NR. 14-3-3s possibly reinforce NR inhibition by magnesium.
FEBS Lett 2000 Sep 01
PMID:Binding to 14-3-3 proteins is not sufficient to inhibit nitrate reductase in spinach leaves. 1103 32

Measurement of nitrite and nitrate, the stable oxidation products of nitric oxide (NO), provides a useful tool to study NO synthesis in vivo and in cell cultures. A simple and rapid fluorometric HPLC method was developed for determination of nitrite through its derivatization with 2,3-diaminonaphthalene (DAN). Nitrite, in standard solution, cell culture medium, or biological samples, readily reacted with DAN under acidic conditions to yield the highly fluorescent 2,3-naphthotriazole (NAT). For analysis of nitrate, it was converted to nitrite by nitrate reductase, followed by the derivatization of nitrite with DAN to form NAT. NAT was separated on a 5-microm reversed-phase C18 column (150X4.6 mm, I.D.) guarded by a 40-microm reversed-phase C18 column (50x4.6 mm, I.D.), and eluted with 15 mM sodium phosphate buffer (pH 7.5) containing 50% methanol (flow-rate, 1.3 ml/min). Fluorescence was monitored with excitation at 375 nm and emission at 415 nm. Mean retention time for NAT was 4.4 min. The fluorescence intensity of NAT was linear with nitrite or nitrate concentrations ranging from 12.5 to 2,000 nM in water, cell culture media, plasma and urine. The detection limit for nitrite and nitrate was 10 pmol/ml. Because NAT is well separated from DAN and other fluorescent components present in biological samples, our HPLC method offers the advantages of high sensitivity and specificity as well as easy automation for quantifying picomole levels of nitrite and nitrate in cell culture medium and biological samples.
J Chromatogr B Biomed Sci Appl 2000 Sep 15
PMID:Rapid determination of nitrite by reversed-phase high-performance liquid chromatography with fluorescence detection. 1107 72

Among the products that are expressed when Mycobacterium tuberculosis undergoes hypoxic shiftdown to nonreplicating persistence (NRP) is the alpha-crystallin chaperone protein homologue (Acr). This expression coincides with the previously reported appearance of a respiratory type of nitrate reductase activity, the increase in glycine dehydrogenase activity, and the production of a unique antigen, URB-1. In a timed sampling study, using a slowly stirred oxygen depletion culture model, we have demonstrated that the hspX mRNA that codes for Acr protein as well as the protein itself is induced just as the bacilli enter the microaerophilic NRP stage 1 (NRP-1). In contrast to the induction observed for hspX mRNA, levels of 16S rRNA, fbpB mRNA (encoding the 85B alpha antigen), and aroB mRNA (encoding dehydroquinate synthase) demonstrate relatively small to no change upon entering NRP-1. Acr protein was shown to be identical to URB-1 by Western analysis with anti-URB-1 antibody. The fact that antibody to Acr is found in a high percentage of tuberculosis patients suggests that the hypoxic shiftdown of tubercle bacilli to the NRP state that occurs in vitro, resulting in production of the alpha-crystallin protein, occurs in vivo as well. Simultaneous abrupt increases in hspX mRNA and Acr protein suggest that Acr protein expression is controlled at the level of transcription.
J Bacteriol 2001 Sep
PMID:Microaerophilic induction of the alpha-crystallin chaperone protein homologue (hspX) mRNA of Mycobacterium tuberculosis. 1151 14

Grain protein content is one of the major determinants of the baking and nutritional quality of wheat. It has previously been reported that the ditelosomic line of wheat (Triticum aestivum L.) CSDT7BL, where the short arm of chromosome 7B is missing, shows a lower grain protein concentration than the normal line, but a similar grain yield. In the present paper the growth and nitrogen (N) metabolism of wheat plants cv. Chinese Spring (CS) and its ditelosomic line CSDT7BL were compared. When plants were grown to maturity in pots with different N supplements, the wild-type line showed a higher grain protein concentration and a lower straw N concentration than the ditelosomic line at every N level analysed, suggesting a deficiency in the N remobilization capacity. When 15-d-old plants were grown in a growth cabinet in pots with sand, and supplied with nutrient solutions of different nitrate concentrations, the ditelosomic line showed no differences in N uptake per unit of root dry weight, nitrate reductase activity, nitrate, total N concentration or free amino acid concentration. However, the ditelosomic line showed a decreased capacity to export amino acids in the phloem under high N, independently of the N source. This deficiency was also observed under dark-induced senescence. The diminished export of amino acids to the phloem was principally caused by a decrease in the export of Glu, Asp, and Gln. It is suggested that the decrease in grain protein concentration in the ditelosomic line is a consequence of defective export in the phloem of these amino acids.
J Exp Bot 2001 Sep
PMID:The export of amino acid in the phloem is altered in wheat plants lacking the short arm of chromosome 7B. 1152 Aug 64

Nitrate or nitrite can be ingested or endogenously produced from nitric oxide. They can cause intoxication and are of general concern for health because they relate to various diseases. Our goal was to study ontogenetic and nutritional effects on the nitrate+nitrite (NOx-) status in cattle, particularly calves. NOx- concentration in blood plasma, cerebrospinal fluid, saliva, and urine was measured based on nitrate conversion by added nitrate reductase to nitrite, which was then determined by the Griess reaction. Concentrations of nitrate were the result of the difference between NOx- and nitrite values. Nitrate in blood plasma, saliva and urine was > or =97% and in cerebrospinal fluid of calves was approximately 35% of NOx-. Preprandial plasma NOx- in calves born after shortened or normal lengths of pregnancy (277 and 290 days) was 470 and 830 micromol/l, respectively, decreased within 4-7 days to 40-60 micromol/l, remained in this range up to 4 months, was < or =5 micromol/l in heifers and no longer measurable in 3-8-year-old cows. Cerebrospinal NOx- in 8-day-old calves was 14 micromol/l and approximately 11-fold lower than in blood plasma. Salivary NOx- decreased postnatally from 600 to 200 micromol/l at 2 days and to 25 micromol/l at 4 weeks. Urinary NOx- excretion decreased from 125 or 16 micromol/l per kg x 24 h in 5-day-old calves to 45 or 8 micromol/kg x 24 h between 10 and 115 days of life and was undetectable in urine of heifers and cows. Feeding neonatal calves no or variable amounts of colostrum, delaying colostrum intake by 24 h after birth or feeding at different feeding intensity had no effect on the NOx- status. In conclusion, the high plasma, salivary and urinary NOx- concentrations especially in newborn calves, ingesting but insignificant amounts of nitrite or nitrate, indicated marked endogenous formation of nitrate, which decreased with age. The high nitrate status may contribute to enhanced susceptibility of young calves to exogenous nitrite+nitrite ingestion.
Comp Biochem Physiol A Mol Integr Physiol 2001 Sep
PMID:High constitutional nitrate status in young cattle. 1154 72


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