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)

Nitric oxide (NO) is known to be synthesized by mammalian cells from L-arginine by a group of NO synthase enzymes. We now show that NO is generated from human skin and propose a different mechanism of production. Whereas enzymatic NO synthesis is inhibited by monomethyl L-arginine, this arginine analog, when infused into the brachial artery at concentrations sufficient to inhibit endothelial NO synthase activity, has little effect on hand skin NO production. Hand skin NO production is increased by topical acidification of the skin surface and greatly increased by the addition of nitrite solutions. We propose that NO generation from skin derives from sweat nitrite (the concentration of which was found to average 3.4 microM in six subjects) due to chemical reduction consequent to the acidic nature of sweat. Sweat contains nitrate in appreciable amounts, and skin commensal bacteria can synthesize nitrate reductase enzyme. Patients on long-term tetracycline antibiotics showed significantly reduced skin NO synthesis, although topical antiseptic and antibiotics had little effect on NO generation in the short-term. We propose that NO generation from skin is dependent on bacterial nitrate reduction to nitrite and subsequent reduction by acidification. We speculate that this has a physiologic role in the inhibition of infection by pathogenic fungi and other susceptible microorganisms and may affect cutaneous T-cell function, keratinocyte differentiation, and skin blood flow.
J Invest Dermatol 1996 Sep
PMID:Nitric oxide is generated on the skin surface by reduction of sweat nitrate. 875 65

Nitrogen-fixing Azotobacter chroococcum cells, but not ammonium- or nitrate-grown cells, exhibited two polypeptide components of 22 and 35 kDa, respectively, that we termed P22 and P35. Bidimensional polyacrylamide gel electrophoresis analysis of preparations from N2-fixing cells that had been transferred to nitrate medium and then incubated for 2 h revealed that P22 had shifted to a more acidic part of the gel while P35 did not change its electrophoretic pattern. Using [32P]orthophosphoric acid it could be demonstrated that the shift in mobility of P22 was due to the phosphorylation of the polypeptide dependent on nitrate (nitrite). The A. chroococcum TR1 strain, which is unable to use nitrate as a nitrogen source and displays activities of nitrogenase, nitrate reductase and nitrite reductase, exhibited both polypeptides. In contrast, P22 and P35 were absent from A. chroococcum MCD1, a mutant strain that cannot assimilate nitrate and lacks the nitrate-reducing enzymatic system. The results suggest that P22 could act as a sensor protein for nitrate in A. chroococcum.
FEBS Lett 1996 Sep 09
PMID:A sensor protein involved in induction of nitrate assimilation in Azotobacter chroococcum. 880 13

The new unstable virescent seedling (vis) allele of a petunia mutant, that has green leaves but white cotyledons with green revertant spots, was used to identify spontaneously occurring haploid petunia lines with active transposable elements. Endogenous transposons were trapped into the single petunia nitrate reductase structural gene (nia) using chlorate selection on haploid protoplasts. In two mutant lines, the dTph1-like transposable element dTph1-3 was inserted at almost the same position but in opposite orientations in the first exon of the nia gene. In a third mutant, a different transposable element was integrated into the fourth exon. This element, called dTph4, is 787 bp long and has 13 bp terminal inverted repeats of which 12 bp are identical to those of dTph1. Insertion of dTph1-3 and dTph4 results in an 8 bp duplication of the target site, as already described for dTph1. In contrast to dTph1-like elements, dTph4 is present at low copy number in the petunia genome. This can facilitate its use for gene tagging in petunia. The dTph1-3 and dTph4 elements excise frequently, as transposon footprints were found in most of the insertion mutants. The data demonstrate that haploid petunia is an excellent system for gene tagging and for the study of transposable elements.
Plant J 1996 Sep
PMID:Insertion mutagenesis and study of transposable elements using a new unstable virescent seedling allele for isolation of haploid petunia lines. 881 67

A new transposable element of tobacco, Slide, was isolated from the tl mutant line, which shows somatic instability, after its transposition into a locus encoding nitrate reductase (NR). The Slide-124 element is 3733 bp long and its coding sequences show similarities with conserved domains of the transposases of Ac, Tam3 and hobo. Excision from the NR locus is detectable in somatic leaf tissues and Slide mobility is triggered by in vitro tissue culture. Slide excision events create footprints similar to those left by Ac and Tam3. Tobacco lines derived from the tl mutant line seem characterized by unmethylated copies of a few members of the highly repetitive Slide family. Slide mobility was monitored in transient expression assays. In wild-type tobacco protoplasts, the complete Slide element, as well as a defective copy, is able to excise. The complete Slide element, but not the defective version, is able to excise in protoplasts of the heterologous species lettuce (Lactuca sativa). These results show that Slide carries the functions required for its own mobility, and represents the first autonomous Ac-like element characterized in Solanaceae species.
Mol Gen Genet 1996 Sep 25
PMID:Molecular and functional characterization of Slide, an Ac-like autonomous transposable element from tobacco. 887 39

Electrostatic properties on the protein surface were examined on the basis of the crystal structure of NADH-cytochrome b5 reductase refined to a crystallographic R factor of 0.223 at 2.1 A resolution and of the other three flavin-dependent reductases. A structural comparison of NADH-cytochrome b5 reductase with the other flavin-dependent reductases, ferredoxin-NADP+ reductase, phthalate dioxygenase reductase, and nitrate reductase, showed that the alpha/beta structure is the common motif for binding pyridine nucleotide. Although the amino acid residues associated with pyridine nucleotide-binding are not conserved, the electrostatic properties and the location of the pyridine nucleotide-binding pockets of NADH-requiring reductases were similar to each other. The electrostatic potential of the surface near the flavin-protruding side (dimethylbenzene end of the flavin ring) of NADH-cytochrome b5 reductase was positive over a wide area while that of the surface near the heme-binding site of cytochrome b5 was negative. This implied that the flavin-protruding side of NADH-cytochrome b5 reductase is suitable for interacting with its electron-transfer partner, cytochrome b5. This positive potential area is conserved among four flavin-dependent reductases. A comparison of the electron-transfer partners of four flavin-dependent reductases showed that there are significant differences in the distribution of electrostatic potential between inter-molecular and inter-domain electron-transfer reactions.
Proteins 1996 Sep
PMID:Electrostatic properties deduced from refined structures of NADH-cytochrome b5 reductase and the other flavin-dependent reductases: pyridine nucleotide-binding and interaction with an electron-transfer partner. 888 Sep 27

The narX, narQ and narL genes of Escherichia coli encode a nitrate-responsive two-component regulatory system that controls the expression of many anaerobic electron-transport- and fermentation-related genes. When nitrate is present, the NarX and NarQ sensor-transmitter proteins function to activate the response-regulator protein, NarL, which in turn binds to its DNA-recognition sites to modulate gene expression. The sensor-transmitter proteins are anchored in the cytoplasmic membrane by two transmembrane domains that are separated by a periplasmic region of approximately 115 amino acids. In this study we report the isolation and characterization of narX* (star) mutants that constitutively activate nitrate reductase (narGHJI) gene expression and repress fumarate reductase (frdABCD) gene expression when no nitrate is provided for the cell. An additional narX mutant was identified that has lost its ability to respond to environmental signals. Each narX defect was caused by a single amino acid substitution within a conserved 17 amino acid sequence, called the 'P-box', in the periplasmic exposed region of the NarX protein. As a result, DNA binding is then 'locked-on' or 'locked-off' to give the observed pattern of gene expression. Diploid analysis of these narX mutants showed that a NarX P-box mutant which conferred a 'locked-on' phenotype was trans dominant over wild-type NarX. Both were also trans dominant over the NarX P-box mutant which conferred a 'locked-off' phenotype. Certain narX P-box mutations, when combined with a narX 'linker' region mutation, were recessive to the NarX linker mutation. Finally, a truncated form of the NarX protein that lacked the periplasmic and membrane regions also showed a 'locked-on' phenotype in vivo. Thus, the periplasmic and membrane domains are essential for signal transduction to NarL. From these findings, we propose that nitrate is detected in the periplasmic space of the cell, and that a signal-transduction event through the cytoplasmic membrane into the interior of the cell modulates the NarX-dependent phosphorylation/dephosphorylation of NarL.
Mol Microbiol 1996 Sep
PMID:Role of the periplasmic domain of the Escherichia coli NarX sensor-transmitter protein in nitrate-dependent signal transduction and gene regulation. 888 62

This work presents the results on two different approaches of RNA-mediated transformation in A. nidulans: a) the receptor strain was an argB2 (III) mutant deficient in arginine (OTCase deficient), and b) the receptor was an A. nidulans mutant defective in nitrate reductase synthesis due to a deletion in the niaD gene (VIII). The analyses of the arg+ and the nia+ retrotransformants allowed an insight on the fate and inheritance of the newly acquired characteristics. The occurrence and the study of Gene Inactivation Mechanism (RIP-like) inactivating the expression of extra copies of genes ectopically scattered over the receptor genome, was a byproduct of this research. Retrotransformants were also used as RNA-donor for a second turn of retrotransformation of the argB and niaD receptor strains. Genetic analyses of the new retrotransformants proved that the retrotransformation ability is kept by the re-extracted RNA when used in a second round of transformation process. This is the best genetic evidence that the newly acquired genetic characteristics were cDNA inserted, precisely transcribed and expressed. These are the first in vivo evidences of genetic information transference mediated by homologous RNA in lower eukaryotes.
Cell Mol Biol (Noisy-le-grand) 1996 Sep
PMID:RNA-mediated transformation in Aspergillus nidulans recovers gene functions lost by deletion or by point mutations. 889 57

The small gene family encoding the chlorophyll a/b-binding proteins of photosystem II (CABII or lhcb) is known to exhibit circadian rhythms of mRNA abundance in Chlamydomonas reinhardtii. In this study we investigated the role of transcription in the phenomenon. We used as reporters Chlamydomonas genes that encode nitrate reductase (NITI) and arylsulfatase (ARS2) transcriptionally fused to sequences upstream of one of the CABII genes (called CABII-1). We found that both reporters exhibited the same circadian rhythm of mRNA abundance in phase, period, and amplitude as does the endogenous CABII-1 gene. We also evaluated the efficacy of arylsulfatase enzymatic activity as a reporter and found that its half-life is too long to make it a useful reporter of rhythmic transcription during a circadian or diurnal cycle. The amount of mRNA synthesis from the CABII-1 gene was examined by in vivo labeling experiments and a circadian rhythm in transcription rate was demonstrated. In vivo labeling also revealed a circadian rhythm of mRNA synthesis for the CABII gene family as a whole. The results from the transcriptional reporter assays together with the in vivo labeling experiments strongly support the conclusion that the biological clock regulates the transcriptional activity of the CABII-I gene, and moreover that regulation at the transcriptional level is the predominant mode by which the clock regulates this gene.
Plant Mol Biol 1996 Sep
PMID:Transcription of CABII is regulated by the biological clock in Chlamydomonas reinhardtii. 891 33

Direct electrochemical studies, utilizing two voltammetric methods-square-wave voltammetry (SWV) and cyclic voltammetry (CV)-have been performed on recombinant forms of the flavin domain of spinach assimilatory nitrate reductase in the presence of NAD+ analogs. The reduction potentials (E degrees ') of the flavin domains have been determined at an edge pyrolytic graphite electrode utilizing MgCl2 as a redox-inactive promoter. Under identical experimental conditions (pH 7.0, 25 degrees C), the two-electron reduction potential for the FAD/FADH2 couple has been determined to be -274 and -257 mV by SWV and CV, respectively. In contrast, the reduction potentials of free FAD have been determined to be -234 and -227 mV by SWV and CV, respectively. The reduction potentials of the complex formed between the FAD prosthetic group in the recombinant flavin domain and various NAD+ analogs have been determined to be as follows: NAD+ (E degrees ' = -192 mV), 5'-ADP ribose (E degrees ' = -199 mV), ADP (E degrees ' = -154 mV), AMP (E degrees ' = -196 mV), adenosine (E degrees ' = -192 mV), adenine (E degrees ' = -220 mV), and NMN (E degrees ' = -208 mV). In contrast to these positive shifts in reduction potential, nicotinamide (E degrees ' = -268 mV) had very little effect on the reduction potential of this flavin complex. Moreover, addition of NAD+ to the FAD prosthetic group in a variety of mutant forms of the recombinant flavin domain resulted in positive shifts in the reduction potential of the complex, although the magnitude of the shifts varied from a minimum of 6 mV obtained for the C240A mutant to a maximum of 79 mV obtained for the C62S mutant. These results represent the first extensive application of direct electrochemistry to examine the redox properties of assimilatory nitrate reductase and indicate that complex formation with NAD+, or various NAD+ analogs, results in a positive shift in the flavin reduction potential, with the magnitude of the shift correlating well with the efficiency of the inhibitor.
Arch Biochem Biophys 1997 Sep 01
PMID:Direct electrochemistry of the flavin domain of assimilatory nitrate reductase: effects of NAD+ and NAD+ analogs. 928 15

The assimilatory nitrate reductase from the phototrophic bacterium Rhodobacter capsulatus has been purified to electrophoretic homogeneity and its molecular and kinetic parameters determined. The native nitrate reductase is a dimer of 144 kDa composed of two subunits of 46 and 95 kDa. The purified enzyme catalyzes the electron transfer from NADH, reduced bromophenol blue or reduced viologens to nitrate. The nitrate reductase contains 1 mol FAD per mole of enzyme and also reduces cytochrome c or dichlorophenol indophenol with NADH as the electron donor. The diaphorase activity is located in the small subunit.
FEBS Lett 1997 Sep 01
PMID:The assimilatory nitrate reductase from the phototrophic bacterium, Rhodobacter capsulatus E1F1, is a flavoprotein. 930 29


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