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Information on EC 1.14.13.39 - nitric-oxide synthase (NADPH) and Organism(s) Rattus norvegicus and UniProt Accession P29476

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IUBMB Comments
The enzyme consists of linked oxygenase and reductase domains. The eukaryotic enzyme binds FAD, FMN, heme (iron protoporphyrin IX) and tetrahydrobiopterin, and its two domains are linked via a regulatory calmodulin-binding domain. Upon calcium-induced calmodulin binding, the reductase and oxygenase domains form a complex, allowing electrons to flow from NADPH via FAD and FMN to the active center. The reductase domain of the enzyme from the bacterium Sorangium cellulosum utilizes a [2Fe-2S] cluster to transfer the electrons from NADPH to the active center. cf. EC 1.14.14.47, nitric-oxide synthase (flavodoxin).
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This record set is specific for:
Rattus norvegicus
UNIPROT: P29476
Word Map
The expected taxonomic range for this enzyme is: Eukaryota, Bacteria
The taxonomic range for the selected organisms is: Rattus norvegicus
Reaction Schemes
hide(Overall reactions are displayed. Show all >>)
Synonyms
bacterial nitric oxide synthase, bacterial nitric-oxide synthase, bNOS, bsNOS, cb-NOS, cytokine inducible NOS, DNOS, e-NOS, EC-NOS, endothelial nitric oxide synthase, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
cb-NOS
248
isoform I, constitutive, from brain
endothelial nitric oxide synthase
endothelial nitric-oxide synthase
248
-
endothelial NOS
248
-
endothelium-derived relaxation factor-forming enzyme
-
-
-
-
endothelium-derived relaxing factor synthase
-
-
-
-
i-NOS
inducible nitric oxide synthase
inducible NO synthase
inducible NOS
287626
isoform
mitochondrial-specific nitric oxide synthase
248
-
n-NOS
NADPH diaphorase
248
-
NADPH-diaphorase
-
-
-
-
neuronal nitric oxide synthase
neuronal nitric-oxide synthase
neuronal NO synthase
287629
-
neuronal NOS
nitric oxide synthase
nitric oxide synthetase
-
-
-
-
nNOSalpha
248
-
NO synthase
NO synthases
287629
-
NOS1
287629
isozyme
NOS3
287632
isozyme
synthetase, nitric oxide
-
-
-
-
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
2 L-arginine + 3 NADPH + 3 H+ + 4 O2 = 2 L-citrulline + 2 nitric oxide + 3 NADP+ + 4 H2O
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
oxidation
-
-
-
-
redox reaction
-
-
-
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reduction
-
-
-
-
SYSTEMATIC NAME
IUBMB Comments
L-arginine,NADPH:oxygen oxidoreductase (nitric-oxide-forming)
The enzyme consists of linked oxygenase and reductase domains. The eukaryotic enzyme binds FAD, FMN, heme (iron protoporphyrin IX) and tetrahydrobiopterin, and its two domains are linked via a regulatory calmodulin-binding domain. Upon calcium-induced calmodulin binding, the reductase and oxygenase domains form a complex, allowing electrons to flow from NADPH via FAD and FMN to the active center. The reductase domain of the enzyme from the bacterium Sorangium cellulosum utilizes a [2Fe-2S] cluster to transfer the electrons from NADPH to the active center. cf. EC 1.14.14.47, nitric-oxide synthase (flavodoxin).
CAS REGISTRY NUMBER
COMMENTARY hide
125978-95-2
-
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
L-arginine + 3 NADPH + 3 H+ + 4 O2
2 L-citrulline + 2 nitric oxide + 3 NADP+ + 4 H2O
show the reaction diagram
L-arginine + NADPH + H+ + O2
citrulline + nitric oxide + NADP+ + H2O
show the reaction diagram
-
-
-
-
?
1-butyl-2-hydroxyguanidine + NADPH + O2
? + NO + NADP+
show the reaction diagram
-
-
-
?
2 L-arginine + 3 NADPH + 4 O2 + 3 H+
2 L-citrulline + 2 NO + 3 NADP+ + 4 H2O
show the reaction diagram
2,6-dichlorophenolindophenol + NADPH + O2
? + NO + NADP+
show the reaction diagram
-
-
-
-
-
2-hydroxy-1-(4-hydroxyphenyl)guanidine + NADPH + O2
? + NO + NADP+
show the reaction diagram
-
-
-
?
2-hydroxy-1-isopropylguanidine + NADPH + O2
? + NO + NADP+
show the reaction diagram
-
-
-
?
L-arginine + NADPH + H+ + O2
citrulline + nitric oxide + NADP+ + H2O
show the reaction diagram
L-arginine + NADPH + H+ + O2
Nomega-hydroxy-L-arginine + NADP+ + H2O
show the reaction diagram
-
first half reaction via intermediate Nomega-hydroxy-L-arginine
-
-
?
L-arginine + NADPH + O2 + tetrahydrobiopterin
citrulline + NO + NADP+ + ?
show the reaction diagram
L-homoarginine + NADPH + O2
?
show the reaction diagram
-
poor substrate
-
-
?
N-hydroxy-L-arginine + NADPH + O2
? + NO + NADP+
show the reaction diagram
Ngamma-hydroxy-L-arginine + NADPH + O2
citrulline + NADP+ + NO
show the reaction diagram
-
substrate is intermediate between reaction 1 and 2 to form citrulline and NO from L-arginine
-
-
ir
nitroblue tetrazolium + NADPH
nitroblue tetrazolium-flavazone + NADP+
show the reaction diagram
-
NADPH-diaphorase reaction
-
?
Nomega-hydroxy-L-arginine + NADPH + H+ + O2
citrulline + nitric oxide + NADP+ + H2O
show the reaction diagram
-
second half reaction via intermediate Nomega-hydroxy-L-arginine
-
-
-
additional information
?
-
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
L-arginine + 3 NADPH + 3 H+ + 4 O2
2 L-citrulline + 2 nitric oxide + 3 NADP+ + 4 H2O
show the reaction diagram
2 L-arginine + 3 NADPH + 4 O2 + 3 H+
2 L-citrulline + 2 NO + 3 NADP+ + 4 H2O
show the reaction diagram
L-arginine + NADPH + H+ + O2
citrulline + nitric oxide + NADP+ + H2O
show the reaction diagram
-
-
-
-
?
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
5,6,7,8-tetrahydro-L-biopterin
Calmodulin
NADPH
(6R)-tetrahydrobiopterin
-
-
2',3'-dialdehyde analogue of NADPH
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activation, can substitute for NADPH at low concentrations, inhibitory at concentrations of 40times the apparent Km-value or after prolonged incubation
5,6,7,8-tetrahydro-L-biopterin
Calmodulin
cytochrome c
-
-
NADPH
tetrahydrobiopterin
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Ca2+
-
low levels of Ca2+ stimulate nNOS activity
Ca2+/calmodulin
calmodulin activates electron transfer from NADPH through three reductase domains to the oxygenase domain, controls constitutive isoforms through regulation of electron transfer between NADPH and heme
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
6-n-propyl-2-thyouracil
-
0.1 mg 6-n-propyl-2-thyouracil decreases nNOS activity to 45% compared to control
A-23187
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high levels of A-23187 inhibit nNOS activity
Ca2+
-
high levels of Ca2+ inhibit nNOS activity
NG-Nitro-L-arginine
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complete inhibition at 1 mM, non-selective NOS inhibitor
Nomega-nitro-L-arginine methyl ester
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1,5,6,7-tetrahydro-2H-azepin-2-imines
-
-
2',3'-dialdehyde of NADPH
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at concentrations of 40times the apparent Km-value or after prolonged incubation, independent of Ca2+/calmodulin, L-arginine or tetrahydrobiopterin, NADPH prevents inhibition, the NADPH-diaphorase activity of the enzyme is less sensitive than the nitric oxide synthase activity
4-(3-amino-propoxy)-1-benzopyran-2-one hydrochloric acid salt
-
IC50: 0.0091 mM
4-(3-dimethylamino-propoxy)-1-benzopyran-2-one hydrochloric acid salt
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IC50: 0.01 mM
4-(3-dimethylamino-propoxy)-1H-quinolin-2-one
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IC50: 0.010 mM
5,6,7,8-tetrahydrobiopterin
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quenches the uncoupled reactions and results in much less reactive oxygen species formation, whereas the presence of redox-incompetent 7,8-dihydrobiopterin demonstrates little quenching effect
6-chloro-4-(3-aminopropoxy)-1-benzopyran-2-one trifluoroacetic acid salt
-
IC50: 0.00056 mM, pharmacokinetic profile
6-chloro-4-(3-dimethylamino-propoxy)-1-benzopyran-2-one hydrochloric acid salt
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IC50: 0.008 mM
6-chloro-4-(3-methylamino-propoxy)-1-benzopyran-2-one trifluoroacetic acid salt
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IC50: 0.00053mM
7-nitroindazole
AR-C102222
-
1,2-dihydro-4-quinazolinamine derivative
AR-C85016
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1,2-dihydro-4-quinazolinamine derivative
AR-R17477
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Ca2+
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preincubation at 37C leads to time-dependent inhibition of the enzyme
Calcineurin
-
-
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Calmidazolium
carbon monoxide
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carbon monoxide down-regulates iNOS activity by reducing its expression level or by inhibiting its activity by converting it to an inactive P420 form, the presence of dithiothreitol, L-Arg, or H4B partially inhibits the iNOSP450 to iNOSP420 conversion, whereas the presence of both L-Arg and 5,6,7,8-tetrahydro-L-biopterin completely prevents the transition
cyanide
-
pretreatment
EDTA
-
brain enzyme
ethylene glycol bis(beta-amino-ethylether)-N,N,N',N'-tetraacetic acid
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i.e. EGTA, complete inhibition of cytosolic enzyme, partial inhibition of particulate enzyme
L-arginine
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L-arginine strongly stimulates oxygen consumption of eNOS and inhibits that of nNOS
L-canavanine
-
liver enzyme, slight inhibition of brain enzyme
N-(4-aminobutyl)-5-chloro-2-naphthalene sulfonamide
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N-(6-Aminohexyl)-1-naphthalene sulfonamide
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NG-Nitro-L-arginine
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complete inhibition at 1 mM, non-selective NOS inhibitor
Ngamma-amino-L-arginine
-
-
Ngamma-iminoethyl-L-ornithine
-
competitive inhibitor
Ngamma-monomethyl-L-arginine
Ngamma-nitro-L-arginine
Ngamma-nitro-L-arginine methyl ester
nitroblue tetrazolium
-
potent non-competitive inhibitor, partially reversible by tetrahydrobiopterin
Nomega-nitro-L-arginine methyl ester
-
nonselective NOS inhibitor
PIN
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human protein enzyme inhibitor, recombinantly expressed in Escherichia coli, the recombinant CREB-binding protein-bound inhibitor protein is purified by calmodulin affinity and inhibits the enzyme to a high extent at 0.001 mM
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S-ethylisothiourea
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inducible NOS inhibitor
Trifluoperazine
additional information
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
A-23187
-
low levels of A-23187 stimulate nNOS activity
Calmodulin
thyroxin
-
0.001 mg thyroxin significantly increases nNOS activity and nNOS protein level to 153% compared to control
-
Ca2+/calmodulin
calmodulin activates electron transfer from NADPH through three reductase domains to the oxygenase domain, controls constitutive isoforms through regulation of electron transfer between NADPH and heme
-
Calmodulin
dithiothreitol
L-arginine
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L-arginine strongly stimulates oxygen consumption of eNOS and inhibits that of nNOS, nonhydrolyzable L-arginine analogues are not stimulatory
additional information
hepatic isozyme iNOS expression is induced in chronic liver cirrhosis early stages
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0008
2',3'-dialdehyde NADPH
-
-
0.008 - 0.0084
dichlorophenolindophenol
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with L-arginine
0.002 - 68.5
L-arginine
0.001 - 0.0029
NADPH
0.00002 - 0.114
tetrahydrobiopterin
additional information
additional information
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.28 - 2.57
NADPH
0.033 - 1.23
nitric oxide
1.08
L-arginine
-
-
0.65
nitroblue tetrazolium
-
NADPH-diaphorase activity
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0012
Ngamma-iminoethyl-L-ornithine
-
-
0.0007 - 0.0065
Ngamma-monomethyl-L-arginine
0.00009 - 0.0004
Ngamma-nitro-L-arginine
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0091
4-(3-amino-propoxy)-1-benzopyran-2-one hydrochloric acid salt
Rattus norvegicus
-
IC50: 0.0091 mM
0.01
4-(3-dimethylamino-propoxy)-1-benzopyran-2-one hydrochloric acid salt
Rattus norvegicus
-
IC50: 0.01 mM
0.01
4-(3-dimethylamino-propoxy)-1H-quinolin-2-one
Rattus norvegicus
-
IC50: 0.010 mM
0.00056
6-chloro-4-(3-aminopropoxy)-1-benzopyran-2-one trifluoroacetic acid salt
Rattus norvegicus
-
IC50: 0.00056 mM, pharmacokinetic profile
0.008
6-chloro-4-(3-dimethylamino-propoxy)-1-benzopyran-2-one hydrochloric acid salt
Rattus norvegicus
-
IC50: 0.008 mM
0.00053
6-chloro-4-(3-methylamino-propoxy)-1-benzopyran-2-one trifluoroacetic acid salt
Rattus norvegicus
-
IC50: 0.00053mM
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.0000000335
-
crude extract
0.000000305
-
crude extract
0.000043
-
crude extract
0.00074
-
crude extract
0.0098
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purified enzyme
0.94 - 0.96
1.9
-
purified enzyme
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.5
-
assay at
additional information
-
pI: 5.6
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
22
-
assay at room temperature
24
-
assay at
30
-
assay at
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
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thoracic, muscle
Manually annotated by BRENDA team
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cerebrum shows higher activity than cerebellum
Manually annotated by BRENDA team
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synaptosomal fraction
Manually annotated by BRENDA team
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estradiol regulates the nitrergic system in the supraoptic and paraventricular hypothalamic nuclei under acute osmotic stress conditions, but the effects specifically depend on the anatomical subregions and different estrogen receptors
Manually annotated by BRENDA team
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NADPH-diaphorase is found in large and small neurons in the sensory, autonomic, and motor nuclei. In contrast to NADPH-diaphorase, nitric oxide synthase in the corresponding nuclei is always present in smaller quantities of mainly smaller neurons. In some nuclei (the nucleus ambiguus, the nucleus of the hypoglossal nerve) containing large numbers of NADPH-diaphorase-positive neurons, nitric oxide synthase immunoreactive cells are particularly rare
Manually annotated by BRENDA team
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islet cells, isoform I
Manually annotated by BRENDA team
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constitutive, soluble form
Manually annotated by BRENDA team
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isoform I
Manually annotated by BRENDA team
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red pulp, eosinophils and neutrophils, isoform II
Manually annotated by BRENDA team
additional information
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
-
total NOS activity is enriched in the intracellular membrane fraction at normal salt diet
Manually annotated by BRENDA team
-
isozyme nNOS
Manually annotated by BRENDA team
-
isozyme iNOS
Manually annotated by BRENDA team
additional information
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
metabolism
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synthesis of the signaling molecule nitric oxide
physiological function
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nitric-oxide synthase (NOS) is required in mammals to generate nitric-oxide for regulating blood pressure, synaptic response, and immune defense
metabolism
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activation of G protein-coupled estrogen receptor in the supraoptic and paraventricular nuclei of the hypothalamus inhibits the phosphorylation of ERK 1/2, which induces a decrease in NADPH-diaphorase expression
physiological function
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
Sequence
NOS1_RAT
1429
0
160559
Swiss-Prot
PDB
SCOP
CATH
UNIPROT
ORGANISM