Information on EC 1.7.2.6 - hydroxylamine dehydrogenase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
1.7.2.6
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RECOMMENDED NAME
GeneOntology No.
hydroxylamine dehydrogenase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
hydroxylamine + 3 ferricytochrome c = nitric oxide + 3 ferrocytochrome c + 3 H+
show the reaction diagram
(2)
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hydroxylamine + H2O + 4 ferricytochrome c = nitrite + 4 ferrocytochrome c + 5 H+
show the reaction diagram
(1)
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
ammonia oxidation I (aerobic)
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Microbial metabolism in diverse environments
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nitrifier denitrification
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Nitrogen metabolism
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nitrate assimilation
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SYSTEMATIC NAME
IUBMB Comments
hydroxylamine:ferricytochrome-c oxidoreductase
The enzymes from the nitrifying bacterium Nitrosomonas europaea [1,4] and the methylotrophic bacterium Methylococcus capsulatus [5] are hemoproteins with seven c-type hemes and one specialized P-460-type heme per subunit. The enzyme converts hydroxylamine to nitrite via an enzyme-bound nitroxyl intermediate [3]. While nitrite is the main product, the enzyme from Nitrosomonas europaea can produce nitric oxide as well [2].
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
strain YNSRA
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Manually annotated by BRENDA team
strain YNSRA
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Manually annotated by BRENDA team
strain ENI-11
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Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
physiological function
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
hydrazine + ferricytochrome c
? + ferrocytochrome c
show the reaction diagram
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-
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?
hydrazine + O2
N2 + H2O
show the reaction diagram
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-
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?
hydrazine + phenazine methosulfate
N2 + reduced phenazine methosulfate
show the reaction diagram
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-
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?
hydroxylamine + 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide + H2O
nitrite + ?
show the reaction diagram
hydroxylamine + ferricyanide
nitrite + ferrocyanide + H+
show the reaction diagram
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more than 90% of added hydroxylamine is oxidized to nitrite within 10 min when 10times as much ferricyanide as hydroxylamine is added
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?
hydroxylamine + ferricytochrome c + H2O
nitrite + ferrocytochrome c + H+
show the reaction diagram
hydroxylamine + ferricytochrome c-554 + H2O
nitrite + ferrocytochrome c-554 + H+
show the reaction diagram
hydroxylamine + ferricytochrome c554
nitric oxide + ferrocytochrome c554 + H+
show the reaction diagram
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-
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?
hydroxylamine + ferricytochrome c554 + H2O
nitrite + ferrocytochrome c554 + H+
show the reaction diagram
hydroxylamine + H2O + 2 ferricyanide
nitrite + 2 ferrocyanide + 5 H+
show the reaction diagram
hydroxylamine + methyl viologen + H+
NH4+ + reduced methyl viologen + H2O
show the reaction diagram
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ir
hydroxylamine + N,N'-bis(carboxymethyl)-N,N'-dinitroso-1,4-phenylenediamine
nitric oxide + ?
show the reaction diagram
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-
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?
hydroxylamine + O2
nitrite + H2O
show the reaction diagram
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phenazine methosulfate, dichlorophenolindophenol, mammalian cytochrome c and 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium chloride can serve as electron acceptors
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?
hydroxylamine + phenazine methosulfate
nitrite + ?
show the reaction diagram
hydroxylamine + phenazine methosulfate + H2O
nitrite + ?
show the reaction diagram
hydroxylamine + Ru(NH3)63++ H+
nitric oxide + Ru(NH3)62+ + H+
show the reaction diagram
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ir
nitrite + methyl viologen + H+
NH4+ + reduced methyl viologen + H2O
show the reaction diagram
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HAO acts like a cytochrome c nitrite reductase, which catalyzes the six-electron reduction of nitrite to NH4+ by reduced methyl viologen
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?
additional information
?
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NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
hydroxylamine + ferricytochrome c + H2O
nitrite + ferrocytochrome c + H+
show the reaction diagram
hydroxylamine + ferricytochrome c554 + H2O
nitrite + ferrocytochrome c554 + H+
show the reaction diagram
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
cytochrome c
purified enzyme shows typical cytochrome-like absorption spectra, but with a peculiar peak at 468 nm in the reduced state that is due to a P460 moiety corresponding to a modified heme c molecule
cytochrome c554
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heme c
additional information
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Ce(NH4)2(NO3)6
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0.1 mM, stimulates the rate of oxidation of NH2OH 2.6-fold and inhibits production of HNO2 by 40%
Mn2+
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0.001 mM, stimulation of NH2OH utilization, stimulation at pH 6 and pH 9 is approximately 40% greater than at pH 7 and pH 8
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2',2'-dipyridyl
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50% inhibition at 0.1 mM
8-hydroxyquinoline
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50% inhibition at 0.2 mM
azide
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50% inhibition at 3.2 mM
Cd2+
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decreases in hydroxylamine oxidoreductase-specific oxygen uptake rate followed by a recovery of hydroxylamine oxidoreductase-specific oxygen uptake rate above steady-state levels do occur upon exposure to Cd2+ concentrations of 0.03 mM and greater
Ce(NH4)2(NO3)6
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0.1 mM, stimulates the rate of oxidation of NH2OH 2.6-fold and inhibits production of HNO2 by 40%
Co(NO3)2
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0.1 mM, 70% inhibition of the rate of nitrite synthesis
cyanide
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active site inhibitor
diethyldithiocarbamate
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inhibition of nitrate production at 0.1 mM
hydrazine
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50% inhibition at 0.1 mM
hydroxyethyl-hydrazine
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methyl-hydrazine
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Mn2+
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complete inhibition of nitrite synthesis at 0.1 mM; complete inhibition of the oxidation of HNO to NO at 0.001 mM, inhibition of HNO2 synthesis is the same at pH 6,7,8 and 9
Na2S
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50% inhibition at 0.3 mM
o-phenanthroline
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50% inhibition at 0.12 mM
phenazine methosulfate
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inhibitory above 0.1 mM
phenyl hydrazine
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N2H3 protects
additional information
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
8-hydroxyquinoline
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1.5fold stimulation at 0.1 mM
cyanide
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2fold stimulation at 1 mM
diethyldithiocarbamate
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stimulation of nitrite production at 0.1 mM
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.017 - 0.2
cytochrome c
0.0009
ferricyanide
pH 7.8, temperature not specified in the publication
0.0011
ferricytochrome c-554
source of substrate Nitrosococcus oceani, pH 7.8, temperature not specified in the publication
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0.003 - 0.005
hydrazine
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
111
ferricyanide
Nitrosococcus oceani
M5DCM0
pH 7.8, temperature not specified in the publication
3.4
ferricytochrome c-554
Nitrosococcus oceani
M5DCM0
source of substrate Nitrosococcus oceani, pH 7.8, temperature not specified in the publication
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383
hydroxylamine
Nitrosomonas europaea
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oxidized
158
nitrite
Nitrosomonas europaea
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additional information
additional information
Nitrosomonas europaea
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overview: effect of pH on rate of reduction of heme c553 of the enzyme by NH2OH, rate constant for reduction of hemes of the enzyme by dithionite at 2C and 19C and by NH2OH or NH2NH2 at 2C
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.01
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crude lysate, in 0.1 M sodium glycine buffer at pH 8.8, temperature not specified in the publication
0.6
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after 60fold purification, in 0.1 M sodium glycine buffer at pH 8.8, temperature not specified in the publication
8.94
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supernatant obtained by centrifugation
11.63
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supernatant obtained by centrifugation
25.6
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supernatant obtained by centrifugation
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
10.5
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rate of reduction of hemes c 553 of the enzyme increases with pH
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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changes in enzyme conformation imposed by changes in solvent, temperature or pressure affect the rates of intramolecular electron transfer from the substrate site to c hemes
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Nitrosomonas europaea (strain ATCC 19718 / CIP 103999 / KCTC 2705 / NBRC 14298)
Nitrosomonas europaea (strain ATCC 19718 / CIP 103999 / KCTC 2705 / NBRC 14298)
Nitrosomonas europaea (strain ATCC 19718 / CIP 103999 / KCTC 2705 / NBRC 14298)
Nitrosomonas europaea (strain ATCC 19718 / CIP 103999 / KCTC 2705 / NBRC 14298)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
11000
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3 * 63000 + 3 * 11000, (alpha,beta)3 subunit structure, SDS-PAGE, 200000 MW enzyme forms a 63000 MW monomer after heme removal, hydroxylamine oxidoreductase probably consists of 3 molecules of monoheme c-type cytochrome with a MW of 11000 and 3 tightly complexed molecules of a catalytically active MW 63000 protein containing 6 c-type hemes and one P-460 heme, SDS-PAGE
16800
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2 * 16800, SDS-PAGE
38900
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gel filtration
64000
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3 * 64000
67000
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3 * 67000
67100
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3 * 67100
68000
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3 * 68000
140000
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active enzyme, gel filtration; SDS-PAGE
175000 - 180000
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SDS-PAGE, gel filtration
180000 - 200000
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SDS-PAGE
180000
x * 180000, SDS-PAGE
189000
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calculated from amino acid sequence
200000
220000
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gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hexamer
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3 * 63000 + 3 * 11000, (alpha,beta)3 subunit structure, SDS-PAGE, 200000 MW enzyme forms a 63000 MW monomer after heme removal, hydroxylamine oxidoreductase probably consists of 3 molecules of monoheme c-type cytochrome with a MW of 11000 and 3 tightly complexed molecules of a catalytically active MW 63000 protein containing 6 c-type hemes and one P-460 heme, SDS-PAGE
homotrimer
oligomer
trimer
additional information
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
hanging drop vapor diffusion method, using 2.4 M ammonium sulfate, 50 mM Tris-HCL, at pH 8.0
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microbatch-under-oil method, using 0.1 M potassium nitrate, 0.1 M MES pH 7.5 and 46% (v/v) PEG 400; using 0.1 M potassium nitrate, 0.1 M MES pH 7.5 and 46% (w/v) PEG 400
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to 2.1 A resolution. Heme P460 contains two covalent cross-links between the porphyrin and a Tyr residue. When purified from source, an unknown physiological HAO binding partner NE1300 is present within the crystal. Protein NE1300 may play a structural role in the ternary complex with cytochrome c554, the physiological electron acceptor of the enzyme
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pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
10.5
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enzyme is permanently inactivated above pH 10.5
394319
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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changes in enzyme conformation imposed by changes in solvent, temperature or pressure affect the rates of intramolecular electron transfer from the substrate site to c hemes
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
relative stable for 72 hours during ammonia starvation
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ORGANIC SOLVENT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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changes in enzyme conformation imposed by changes in solvent, temperature or pressure affect the rates of intramolecular electron transfer from the substrate site to c hemes
OXIDATION STABILITY
ORGANISM
UNIPROT
LITERATURE
on exposure to air, the 100% reduced enzyme sample is oxidized within about 1 min
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394321
the enzyme is inactivated by incubation of 1 mM enzyme with a 5fold excess of peroxide for 1 h. The enzyme is also inactivated in the presence of H2O2
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719635
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ammonium sulfate precipitation
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ammonium sulfate precipitation, DEAE cellulose column chromatography, gel filtration
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ammonium sulfate precipitation, DEAE-cellulose column chromatography, and Sephadex G-200 gel filtration
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ammonium sulfate precipitation, octyl-Sepharose column chromatography, and Sephacryl S-200 gel filtration
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ammonium sulfate precipitation, Sepharose CL-6B column chromatography, DEAE-Sephacel column chromatography, and Sephadex G-100 gel filtration; ammonium sulfate precipitation, Sepharose CL-6B column chromatography, DEAE-Sephacel gel filtration, and Sephadex G100 gel filtration
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ammonium sulfate precipitation, Sepharose CL-6B column chromatography, DEAE-Sephacel gel filtration, octyl Sepharose column chromatography, and Sephadex G-100 gel filtration
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
cloned and expressed in Pseudomonas putida
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expressed in Escherichia coli DH5alpha cells
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EXPRESSION
ORGANISM
UNIPROT
LITERATURE
mRNA expression is upregulated in the presence of NH4+
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mRNA expression is upregulated in the presence of NH4+; there is some increase in the enzyme level only after 4 h incubation of starved Nitrosomonas europaea cells with 2 mM ammonia
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the enzyme level remains highly stable during ammonia starvation. The enzyme level shows no significant change when the cells are incubated with 0.2 mM ammonia for 60 min at 4C or 26C
there is some increase in the enzyme level only after 4 h incubation of starved Nitrosomonas europaea cells with 2 mM ammonia
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis
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use of hydroxylamine oxidoreductase to differentiate Nitrosomonas spp. and to identify autotrophic ammonia oxidizing bacteria. The ensembled use of single strand confirmation polymorphism and HAO enzyme zymogram in fingerprinting AOB provide better resolution and evenness, contributing significantly in autotrophic ammonia oxidizing bacteria diversity studies