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Information on EC 1.7.2.2 - nitrite reductase (cytochrome; ammonia-forming) and Organism(s) Escherichia coli and UniProt Accession P0ABK9

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EC Tree
IUBMB Comments
Found as a multiheme cytochrome in many bacteria. The enzyme from Escherichia coli contains five hemes c and requires Ca2+. It also reduces nitric oxide and hydroxylamine to ammonia, and sulfite to sulfide.
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This record set is specific for:
Escherichia coli
UNIPROT: P0ABK9
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Word Map
The taxonomic range for the selected organisms is: Escherichia coli
The enzyme appears in selected viruses and cellular organisms
Reaction Schemes
Synonyms
cytochrome c-552, cytochrome c nitrite reductase, ccnir, tvnir, nrfha, octahaem cytochrome c nitrite reductase, hexaheme c-type cytochrome, cytochrome c nitrite reductase complex, sco2488, nrfa2nrfh complex, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
cytochrome c nitrite reductase
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cytochrome c-552
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cytochrome c nitrite reductase
multiheme nitrite reductase
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-
-
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
redox reaction
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-
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oxidation
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-
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reduction
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SYSTEMATIC NAME
IUBMB Comments
ammonia:ferricytochrome-c oxidoreductase
Found as a multiheme cytochrome in many bacteria. The enzyme from Escherichia coli contains five hemes c and requires Ca2+. It also reduces nitric oxide and hydroxylamine to ammonia, and sulfite to sulfide.
CAS REGISTRY NUMBER
COMMENTARY hide
37256-41-0
-
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
hydroxylamine + reduced benzyl viologen
? + oxidized benzyl viologen
show the reaction diagram
-
-
-
?
nitrite + ferrocytochrome c + H+
NH3 + H2O + ferricytochrome c
show the reaction diagram
nitrite + reduced methyl viologen
NH3 + H2O + oxidized methyl viologen
show the reaction diagram
hydroxylamine + ferrocytochrome c
? + ferricytochrome c
show the reaction diagram
-
-
-
-
?
hydroxylamine + reduced methyl viologen
?
show the reaction diagram
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low activity
-
-
?
hydroxylammonium + ferrocytochrome c
NH3 + ferricytochrome c + H2O
show the reaction diagram
-
-
-
-
?
nitric oxide + ferrocytochrome c
NH3 + ferricytochrome c + H2O
show the reaction diagram
-
-
-
-
?
nitrite + 6 ferrocytochrome c + 7 H+
NH3 + 2 H2O + 6 ferricytochrome c
show the reaction diagram
-
-
-
-
?
nitrite + ferrocytochrome c
NH3 + ferricytochrome c + H2O
show the reaction diagram
-
-
-
-
?
nitrite + ferrocytochrome c + 6 H+
NH3 + ferricytochrome c + 2 H2O
show the reaction diagram
-
-
-
-
?
nitrite + reduced methyl viologen
NH3 + H2O + oxidized methyl viologen
show the reaction diagram
-
-
-
-
?
NO + 5 ferrocytochrome c + 5 H+
NH3 + H2O + 5 ferricytochrome c
show the reaction diagram
NO radical + reduced methyl viologen
?
show the reaction diagram
-
-
-
-
?
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
nitrite + ferrocytochrome c + H+
NH3 + H2O + ferricytochrome c
show the reaction diagram
nitrite + 6 ferrocytochrome c + 7 H+
NH3 + 2 H2O + 6 ferricytochrome c
show the reaction diagram
-
-
-
-
?
NO + 5 ferrocytochrome c + 5 H+
NH3 + H2O + 5 ferricytochrome c
show the reaction diagram
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important role for the enzyme in nitric oxide management in oxygen-limited environments. Nitric oxide is a key element in host defense against invasive pathogens
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?
additional information
?
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NrfA from Escherichia coli has a well established role in the respiratory reduction of nitrite to ammonium, it might also participate in NO radical detoxification, detoxifying exogenously generated NO radical encountered during invasion of a human host, overview
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-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
methyl viologen
artificial cofactor, zinc-reduced or dithionite-reduced
methyl viologen
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used as the electron donor, reduced with dithionite, artificial cofactor
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Fe3+
active site heme Fe(III) iron, the NrfA active site consists of a hexacoordinate high-spin heme with a lysine ligand on the proximal side and water/hydroxide or substrate on the distal side. There are four further highly conserved active site residues including a Q263 positioned near the heme iron for which the side chain, unusually, coordinates a conserved, essential calcium ion, overview
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Zn2+
EDTA protects NrfA from zinc inhibition, zinc inhibition kinetics in presence or absence of EDTA, overview
azide
CN-
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cyanide is a potent inhibitor of NrfA nitrite and hydroxylamine reductase activities
Cyanate
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cyanide
nitrate
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thiocyanate
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effective inhibitor
additional information
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
30 - 170
hydroxylamine
0.033 - 0.413
nitrite
60.4
hydroxylamine
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pH 7.0, 20°C, reduction
0.00025 - 0.012
nitrite
0.3
NO radical
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pH 7.0, 20°C, reduction
additional information
additional information
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
3100 - 3160
hydroxylamine
1350
nitrite
wild-type, pH 7.0, temperature not specified in the publication
390
nitric oxide
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turnover rate in intact cell
700
nitrite
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pH 7.0, 20°C, reduction
840
NO radical
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pH 7.0, 20°C, reduction
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25
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nitrite reduction
additional information
-
quantitation of steady-state NrfA reduction of NO radical, nitrite, and hydroxylamine
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7
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assay at
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
-
pH dependence of NrfA at 4-25°C, overview
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
20
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assay at
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4 - 25
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NrfA activity increases when the temperature is raised from 4°C to 25°C
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homodimer
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CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
mutant H264N, to 2.15 A resolution. Homodimeric protein, the constellation of His/His and Lys-coordinated c-hemes is indistinguishable from those of the native enzyme
purified NrfA, hanging drop vapor diffusion method, five conditions that result in several different crystal packing motifs, e.g. 0.001 ml of protein solution containing 10 mg/ml of NrfA mixed with 0.001 ml of mother liquor containing 100 mM HEPES, pH 7.5, 20% PEG 10000 and incubated in hanging drop trays with a 1 ml well solution at 4°C, 5-14 days, other conditions, overview. X-ray diffraction structure determination and analysis at 1.7-2.5 A resolution
purified recombinant wild-type and mutant Q263E, 10 mg/ml protein, under aerobic conditions by the vapor diffusion hanging drop method using 20% v/v PEG 10000 in 100 mM Na-HEPES, pH 7.5, 20% ethylene glycol as cryoprotectant, X-ray diffraction structure determination and analysis at 1.74 A and 2.04 A resolution, respectively, molecular replacement
vapour diffusion method. Crystals belong to space group P2(1)2(1)2(1) with apparent cell parameters of a = 81.47 A, b = 90.84 and c = 294.87 A and contain four molecules of NrfA per asymmetric unit
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PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
H264N
mutant is unable to catalyze nitrite reduction but able to reduce hydroxylamine. The mutant simultaneously binds nitrite and electrons at the catalytic heme
Q263E
site-directed mutagenesis, the mutation leads to introduction of a negative charge into the vicinity of the active site heme, and the mutant shows reduced activity compared to the wild-type enzyme. The high spin state of the active site to be preserved, indicating that a water/hydroxide molecule is still coordinated to the heme in the resting state of the enzyme
additional information
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anaerobically grown Escherichia coli nrf mutants are more sensitive to NO radicals than the parent strain
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant wild-type and mutant enzymes from Escherichia coli strain JCB4083a by ammonium sulfate fractionation and anion exchange chromatography
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli
gene nrfA, expression of wild-type and mutant enzymes in Escherichia coli strain JCB4083a
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Bamford, V.A.; Angove, H.C.; Seward, H.E.; Thomson, A.J.; Cole, J.A.; Butt, J.N.; Hemmings, A.M.; Richardson, D.J.
Structure and spectroscopy of the periplasmic cytochrome c nitrite reductase from Escherichia coli
Biochemistry
41
2921-2931
2002
Escherichia coli
Manually annotated by BRENDA team
Gwyer, J.D.; Richardson, D.J.; Butt, J.N.
Resolving complexity in the interactions of redox enzymes and their inhibitors: contrasting mechanisms for the inhibition of a cytochrome c nitrite reductase revealed by protein film voltammetry
Biochemistry
43
15086-15094
2004
Escherichia coli
Manually annotated by BRENDA team
Gwyer, J.D.; Angove, H.C.; Richardson, D.J.; Butt, J.N.
Redox-triggered events in cytochrome c nitrite reductase
Bioelectrochemistry
63
43-47
2004
Escherichia coli
Manually annotated by BRENDA team
Poock, S.R.; Leach, E.R.; Moir, J.W.; Cole, J.A.; Richardson, D.J.
Respiratory detoxification of nitric oxide by the cytochrome c nitrite reductase of Escherichia coli
J. Biol. Chem.
277
23664-23669
2002
Escherichia coli
Manually annotated by BRENDA team
Burlat, B.; Gwyer, J.D.; Poock, S.; Clarke, T.; Cole, J.A.; Hemmings, A.M.; Cheesman, M.R.; Butt, J.N.; Richardson, D.J.
Cytochrome c nitrite reductase: from structural to physicochemical analysis
Biochem. Soc. Trans.
33
137-140
2005
Escherichia coli
Manually annotated by BRENDA team
Gwyer, J.D.; Richardson, D.J.; Butt, J.N.
Inhibiting Escherichia coli cytochrome c nitrite reductase: voltammetry reveals an enzyme equipped for action despite the chemical challenges it may face in vivo
Biochem. Soc. Trans.
34
133-135
2006
Escherichia coli
Manually annotated by BRENDA team
Clarke, T.A.; Kemp, G.L.; Van Wonderen, J.H.; Doyle, R.M.; Cole, J.A.; Tovell, N.; Cheesman, M.R.; Butt, J.N.; Richardson, D.J.; Hemmings, A.M.
Role of a conserved glutamine residue in tuning the catalytic activity of Escherichia coli cytochrome c nitrite reductase
Biochemistry
47
3789-3799
2008
Escherichia coli (P0ABK9), Escherichia coli
Manually annotated by BRENDA team
van Wonderen, J.H.; Burlat, B.; Richardson, D.J.; Cheesman, M.R.; Butt, J.N.
The nitric oxide reductase activity of cytochrome c nitrite reductase from Escherichia coli
J. Biol. Chem.
283
9587-9594
2008
Escherichia coli
Manually annotated by BRENDA team
Clarke, T.A.; Mills, P.C.; Poock, S.R.; Butt, J.N.; Cheesman, M.R.; Cole, J.A.; Hinton, J.C.; Hemmings, A.M.; Kemp, G.; Soederberg, C.A.; Spiro, S.; Van Wonderen, J.; Richardson, D.J.
Escherichia coli cytochrome c nitrite reductase NrfA
Methods Enzymol.
437
63-77
2008
Escherichia coli (P0ABK9), Escherichia coli
Manually annotated by BRENDA team
Lockwood, C.W.; Burlat, B.; Cheesman, M.R.; Kern, M.; Simon, J.; Clarke, T.A.; Richardson, D.J.; Butt, J.N.
Resolution of key roles for the distal pocket histidine in cytochrome C nitrite reductases
J. Am. Chem. Soc.
137
3059-3068
2015
Escherichia coli (P0ABK9), Wolinella succinogenes (Q9S1E5), Wolinella succinogenes DSM 1740 (Q9S1E5)
Manually annotated by BRENDA team