random order rapid-equilibrium mechanism, two functional parts: 1. heat labile, FAD and haemoprotein containing, binds NADPH and transfers electrons from NADPH via FAD and perhaps cytochrome b to acceptors such as cytochrome c, 2. heat stable, molybdenum containing, accepts electrons from reduced viologen dyes and transfers them to nitrate
multicenter redox enzyme. Ser920, Arg921 and Arg932 are suggested to be the key enzymes to investigate for a role in determining pyridine nucleotide specificity. Arg932 may be playing a role in binding the adenine ring of NADPH
active site formation of eukaryotic nitrate reductase is an autonomous process intrinsically tied to nitrate reductase dimerization, molybdenum cofactor-dependent enzyme maturation, overview
enzyme is essential for reduction of nitrate to ammonia, under oxic conditions the enzyme is involved in nitrate assimilation, under anoxic conditions the enzyme is used for dissimilatory nitrate reduction, transcription regulation mechanism via ammonium, nitrate, and O2 concentrations, overview
catalyzes the NADPH-linked reduction of ferricyanide and 2,6-dichlorophenolindophenol, chlorate- and bromate-dependent NADPH oxidation, and FMNH-linked nitrate reduction
key enzyme in the assimilation path of nitrate to ammonium. Crude extracts possess endogenous NADPH regenerating systems capable of providing reducing equivalents for effective nitrate reduction in vitro
key enzyme in the assimilation path of nitrate to ammonium. Crude extracts possess endogenous NADPH regenerating systems capable of providing reducing equivalents for effective nitrate reduction in vitro
enzyme expression is regulated by different inorganic and organic nitrogen sources, e.g. nitrate, ammonium, urea and glutamate, regulatory mechanism of nitrate aquisition in ectomycchorizae, overview
NADPH-dependent cytochrome c reducing activity by the holo-enzyme is determined with FAD and NADPH spectroscopically at 550 nm and 340 nm. Apo-nitrate reductase has a marginally lower, about 10% reduced cytochrome c reducing activity, which correlates to its 15% reduced heme content
enzyme expression is regulated by different inorganic and organic nitrogen sources, e.g. nitrate, ammonium, urea and glutamate, regulatory mechanism of nitrate aquisition in ectomycchorizae, overview
enzyme is essential for reduction of nitrate to ammonia, under oxic conditions the enzyme is involved in nitrate assimilation, under anoxic conditions the enzyme is used for dissimilatory nitrate reduction, transcription regulation mechanism via ammonium, nitrate, and O2 concentrations, overview
key enzyme in the assimilation path of nitrate to ammonium. Crude extracts possess endogenous NADPH regenerating systems capable of providing reducing equivalents for effective nitrate reduction in vitro
key enzyme in the assimilation path of nitrate to ammonium. Crude extracts possess endogenous NADPH regenerating systems capable of providing reducing equivalents for effective nitrate reduction in vitro
i.e Moco/MPT, binding of molybdenum cofactor to apo-nitrate reductase is independent from other prosthetic groups, molybdenum cofactor-dependent enzyme maturation, overview. Reconstitution of Moco-free nitrate reductase with various amounts of purified Moco carrier protein
cofactor is necessary and sufficient to induce dimer formation. The molybdenum center of nitrate reductase reconstituted in vitro from apo-enzyme and cofactor shows an EPR spectrum identical to holo-enzyme. Insertion of this cofactor into the enzyme occurs independent from the insertion of any other NR redox cofactor
i.e Moco/MPT, binding of molybdenum cofactor to apo-nitrate reductase is independent from other prosthetic groups, molybdenum cofactor-dependent enzyme maturation, overview. Reconstitution of Moco-free nitrate reductase with various amounts of purified Moco carrier protein
represses the intracellular NO3- assimilation, which is required for the induction and maintenace of NADPH nitrate reductase, enzyme is regulated by NH4+ at the transcriptional level
exogenous ammonium represses enzyme expression under oxic conditions in presence or absence of nitrate, while under anoxic conditions the enzyme is expressed even in the presence of ammonium, promotor activity study
intracellularly accumulated nitrite can inhibit nitrate uptake, addition of nitrite at concentrations above 5 mM is toxic and causes growth retardation
exogenous nitrate induces the enzyme expression, can be repressed by ammonium under aerobic conditions, but not under anaerobic conditions, promotor activity study
the enzymatic activity is increased in root ectomycchorizae in symbiosis with the fungus Tuber borchii, while the expression of the endogenous enzyme is decreased
aggregate of two different polypeptide chains: one responsible for transport of electrons fom NADPH to FAD or cytochrome c, nit-1 enzyme. The other transfers electrons from FAD via molybdenum to nitrate, nit-3 enzyme; nit-1 mutant enzyme is the apoprotein of nitrate reductase
the apo-enzyme dissociates completely into monomers, the ratio between monomeric and dimeric apo-NR does not change significantly upon a 20fold dilution. Active site formation of eukaryotic nitrate reductase is an autonomous process intrinsically tied to nitrate reductase dimerization, molybdenum cofactor-dependent enzyme maturation, overview. Enzyme domain structure, overview
-10°C, 50 mM sodium phosphate, pH 6.9, 30% glycerol, 0.5 mM EDTA, 0.5 mM dithiothreitol, 0.5 mM phenylmethylsulfonylfluoride, 0.1 mM FAD, stable for at least 6 months
5°C, urea-treated, 0.1 M potassium phosphate buffer, pH 7.3, 5 mM EDTA, 2 mM dithioerythritol, 0.5 mg/ml or more bovine serum albumin, stable for 18 h without loss of activity
partially purification of wild-type and nit-3 mutant by streptomycin sulfate precipitation and ammonium sulfate fractionation, of nit-1 mutant by protamine sulfate fractionation, ammonium sulfate precipitation and gel filtration
quick purification by immunoprecipitation with monospecific anti-nitrate reductase serum, stabilization during purification and protection of proteolytic cleavage by addition of phenylmethylsulphonyl fluoride
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
construction of a fusion gene comprising gene the promotor of gene niaD and the Escherichia coli beta-glucuronidase gene, i.e. GUS, for analysis of promotor activity in Aspergillus nidulans
gene nar1, DNA and amino acid sequence determination and analysis, expression study, gene nar1 is part of the nitrate assimilation gene cluster together with genes encoding a high-affinity nitrate transporter and a nitrite reductase, the intergenic region could also act as bidirectional promotor, organisation of coding sequences
site-directed mutagenesis, CD spectroscopy shows no negative effects of the introduced mutations on protein secondary structure in comparison to the wild-type protein, but the mutant contains no heme, while the FAD binding ability is not significantly disturbed
important for the enzyme's interaction with the pyridine nucleotide substrates. Mutant retains ~2% of the NADPH activity of the wild type while it has an increased NADH activity, ~15% higher. It is concluded that Ser920 is a ligand involved in binding the 2' phosphate of NADPH in the wild type enzyme
greatest decrease in NADPH activity of all created mutants, shows that Arg932 is a residue interacting with the pyridine nucleotide coenzyme electron donors and that Ser920 and Arg932 have effects on substrate binding and catalytic activity. Both residues may be ligands to the 2' phosphate of NADPH in the wild type cyt b reductase fragment of nitrate reductase
studies of temperature-sensitive mutations, niaD gene: mutation leads to loss of a 4.5-S cytochrome-c reductase activity, which is a subunit of nitrate reductase. It is suggested that neither the product of the cnxE nor the cnyF genes form part of the nitrate reductase molecule, but some catalytic role in cofactor formation, niaD and cnxH seem to be structural genes
studies of temperature-sensitive mutations, niaD gene: mutation leads to loss of a 4.5-S cytochrome-c reductase activity, which is a subunit of nitrate reductase. It is suggested that neither the product of the cnxE nor the cnyF genes form part of the nitrate reductase molecule, but some catalytic role in cofactor formation, niaD and cnxH seem to be structural genes
the purified native enzyme is successfully used for synthesis of silver nanoparticles in an NADPH-dependent manner using gelatin as a capping agent, analysis by X-ray diffraction, dynamic light scattering spectroscopy, and transmission and scanning electron microscopy, overview. 0.2 M Phosphate buffer, pH 7.2, 1 mM silver nitrate as the enzyme substrate, 0.1 mg gelatin as a capping agent, 1 mM 4-hydroxyquinoline as an electron carrier, 1 mM NADPH as enzyme cofactor, and 0.1 mg of purified fungal nitrate reductase are incubated at 25°C for 5 h. The stable nonaggregating nanoparticles are spherical in shape with an average size of 50 nm and a zeta potential of -34.3. The synthesized nanoparticles show a strong growth inhibitory antimicrobial activity against all tested human pathogenic fungi and bacteria, overview
the purified native enzyme is successfully used for synthesis of silver nanoparticles in an NADPH-dependent manner using gelatin as a capping agent, analysis by X-ray diffraction, dynamic light scattering spectroscopy, and transmission and scanning electron microscopy, overview. 0.2 M Phosphate buffer, pH 7.2, 1 mM silver nitrate as the enzyme substrate, 0.1 mg gelatin as a capping agent, 1 mM 4-hydroxyquinoline as an electron carrier, 1 mM NADPH as enzyme cofactor, and 0.1 mg of purified fungal nitrate reductase are incubated at 25°C for 5 h. The stable nonaggregating nanoparticles are spherical in shape with an average size of 50 nm and a zeta potential of -34.3. The synthesized nanoparticles show a strong growth inhibitory antimicrobial activity against all tested human pathogenic fungi and bacteria, overview
several mutations of recombinant cyt b reductase fragment of nitrate reductase in the region Ser920, Arg921 and Arg932 are created. Conversion from NADPH-specific to virtually NADH-specific cyt b reductase fragment of nitrate reductase
nit-1 mutant: lacks all activities except FAD-dependent NADPH:cytochrome c reductase activity,nit-2 mutant: reduced FAD:- and reduced methyl viologen:nitrate reductase activities but lacks the other two activities
the purified native enzyme is successfully used for synthesis of silver nanoparticles in an NADPH-dependent manner using gelatin as a capping agent, analysis by X-ray diffraction, dynamic light scattering spectroscopy, and transmission and scanning electron microscopy, overview. The stable nonaggregating nanoparticles are spherical in shape with an average size of 50 nm and a zeta potential of -34.3. The synthesized nanoparticles show a strong growth inhibitory antimicrobial activity against all tested human pathogenic fungi and bacteria, overview
the purified native enzyme is successfully used for synthesis of silver nanoparticles in an NADPH-dependent manner using gelatin as a capping agent, analysis by X-ray diffraction, dynamic light scattering spectroscopy, and transmission and scanning electron microscopy, overview. The stable nonaggregating nanoparticles are spherical in shape with an average size of 50 nm and a zeta potential of -34.3. The synthesized nanoparticles show a strong growth inhibitory antimicrobial activity against all tested human pathogenic fungi and bacteria, overview
the purified native enzyme is successfully used for synthesis of silver nanoparticles in an NADPH-dependent manner using gelatin as a capping agent, analysis by X-ray diffraction, dynamic light scattering spectroscopy, and transmission and scanning electron microscopy, overview. The stable nonaggregating nanoparticles are spherical in shape with an average size of 50 nm and a zeta potential of -34.3. The synthesized nanoparticles show a strong growth inhibitory antimicrobial activity against all tested human pathogenic fungi and bacteria, overview
the purified native enzyme is successfully used for synthesis of silver nanoparticles in an NADPH-dependent manner using gelatin as a capping agent, analysis by X-ray diffraction, dynamic light scattering spectroscopy, and transmission and scanning electron microscopy, overview. The stable nonaggregating nanoparticles are spherical in shape with an average size of 50 nm and a zeta potential of -34.3. The synthesized nanoparticles show a strong growth inhibitory antimicrobial activity against all tested human pathogenic fungi and bacteria, overview
Purification and characterization of homogeneous assimilatory reduced nicotinamide adenine dinucleotide phosphate-nitrate reductase from Neurospora crassa
Engineering of pyridine nucleotide specificity of nitrate reductase: mutagenesis of recombinant cytochrome b reductase fragment of Neurospora crassa NADPH:nitrate reductase
Purification and properties of the nitrate reductase isolated from Neurospora crassa mutant nit-3. Kinetics, molecular weight determination, and cytochrome involvement
Characterisation and expression analysis of a nitrate transporter and nitrite reductase genes, two members of a gene cluster for nitrate assimilation from the symbiotic basidiomycete Hebeloma cylindrosporum
Gholami-Shabani, M.; Akbarzadeh, A.; Norouzian, D.; Amini, A.; Gholami-Shabani, Z.; Imani, A.; Chiani, M.; Riazi, G.; Shams-Ghahfarokhi, M.; Razzaghi-Abyaneh, M.
Antimicrobial activity and physical characterization of silver nanoparticles green synthesized using nitrate reductase from Fusarium oxysporum