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Information on EC 1.7.2.2 - nitrite reductase (cytochrome; ammonia-forming) and Organism(s) Thioalkalivibrio nitratireducens and UniProt Accession L0DSL2

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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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Thioalkalivibrio nitratireducens
UNIPROT: L0DSL2
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The taxonomic range for the selected organisms is: Thioalkalivibrio nitratireducens
The enzyme appears in selected viruses and cellular organisms
Reaction Schemes
hide
NH3
+
2
H2O
+
6
ferricytochrome c
=
nitrite
+
6
ferrocytochrome c
+
7
H+
+
2
+
6
=
+
6
+
7
Synonyms
cytochrome c-552, cytochrome c nitrite reductase, ccnir, tvnir, nrfha, octahaem cytochrome c nitrite reductase, cytochrome c nitrite reductase complex, hexaheme c-type cytochrome, multiheme cytochrome c nitrite reductase, sco2488, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
cytochrome c nitrite reductase
Q5F2I3
-
octahaem cytochrome c nitrite reductase
Q5F2I3
-
cytochrome c nitrite reductase
multiheme nitrite reductase
-
-
-
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
redox reaction
-
-
-
-
oxidation
-
-
-
-
reduction
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
nitrite + acceptor
NH3 + oxidized acceptor + H2O
show the reaction diagram
Q5F2I3
-
-
-
?
nitrite + reduced methyl viologen
NH3 + H2O + oxidized methyl viologen
show the reaction diagram
Q5F2I3
-
-
-
?
nitrite + reduced methyl viologen
NH3 + oxidized methyl viologen + H2O
show the reaction diagram
Q5F2I3
-
-
-
?
sulfite + reduced methyl viologen
H2S + oxidized methyl viologen + H2O
show the reaction diagram
Q5F2I3
-
-
-
?
sulfite + reduced methyl viologen + H+
H2S + oxidized methyl viologen + H2O
show the reaction diagram
Q5F2I3
-
-
-
?
hydroxylamine + ferrocytochrome c
NH3 + ferricytochrome c + H2O
show the reaction diagram
nitrite + ferrocytochrome c
NH3 + ferricytochrome c + H2O
show the reaction diagram
-
-
-
-
?
sulfite + ?
sulfide + ?
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 + acceptor
NH3 + oxidized acceptor + H2O
show the reaction diagram
Q5F2I3
-
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Ferredoxin
Q5F2I3
[4Fe-4S]-ferredoxin cofactor
-
heme
Q5F2I3
an octaheme cytochrome c nitrite reductase
methyl viologen
Q5F2I3
used as the electron donor, reduced with dithionite, artificial cofactor
heme
-
7.5 units heme per subunit
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Iron
Q5F2I3
[4Fe-4S]-ferredoxin cofactor
Ca2+
-
1-3 mol per subunit
Fe
-
8-11 mol per subunit
Iron
-
contains 16 Fe atoms
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
cyanide
-
strong inhibitor
phosphate
-
weak inhibitor
sulfate
-
weak inhibitor
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
16.4
hydroxylamine
-
in 0.1 M K+-phosphate buffer, at pH 7.0 and 30°C
16.7
nitrite
-
in 0.1 M K+-phosphate buffer, at pH 7.0 and 30°C
60
sulfite
-
in 0.1 M K+-phosphate buffer, at pH 7.0 and 30°C
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1195 - 2860
nitrite
0.023 - 0.039
sulfite
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
150
Q5F2I3
purified native enzyme
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7 - 9
Q5F2I3
the sulfite reductase activity of NiR has a maximum value at neutral pH, the sulfite reductase activity of NiR decreases with increasing pH and is absent at pH 9.0, the activity measured at pH 7.8 is 20% of the activity measured at pH 7.0
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
380000 - 390000
Q5F2I3
gel filtration
64000
-
6 * 64000, calculated from sequence analysis and spectral properties
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hexamer
Q5F2I3
determination of the oligomeric composition of NiR by small-angle X-ray scattering, overview
hexamer
-
6 * 64000, calculated from sequence analysis and spectral properties
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
alone or in complex with nitrite and cyanide, hanging drop vapor diffusion method, using 0.2 M trisodium citrate dihydrate, 0.1 M Tris-HCl pH 8.5 and 30% (v/v) PEG 400
Q5F2I3
apoenzyme and its complexes with the substrate nitrite and the inhibitor azide. The subunit of NiR consists of the N-terminal domain which has an unique fold and contains three hemes and the catalytic C-terminal domain which hosts the remaining five hemes. The complete set of eight hemes forms a spatial pattern characteristic of other multiheme proteins, including structurally characterized octaheme cytochromes. The catalytic machinery comprises the lysine residue at the proximal position of the catalytic heme, the catalytic triad of tyrosine, histidine, and arginine at the distal side, channels for the substrate and product transport with a characteristic gradient of electrostatic potential, and, two conserved Ca2+-binding sites. In addition, the enzyme has a covalent bond between the catalytic tyrosine and the adjacent cysteine and an unusual topography of the product channels that open into the void interior space of the protein hexamer
Q5F2I3
modified form of the enzyme that contains an additional covalent bond between residues Tyr303 and Gln360 in complex with phosphate to 1.45 A resolution, and with sulfite to 1.8 A resolution, structure of unmodified enzyme in complex with nitrite to 1.83 A resolution. Structure reveal the presence of a covalent bond between the CE2 atom of the catalytic Tyr303 and the S atom of Cys305, which might be responsible for the higher nitrite reductase activity. The formation of the second covalent bond by Tyr303 leads to a decrease in both the nitrite and sulfite reductase activities of the enzyme. Tyr303 is located at the exit from the putative proton-transport channel to the active site
Q5F2I3
diffraction data sets with increasing absorbed doses. The structures reveal gradual changes associated with the reduction of the catalytic hemes by X-rays. The conversion of the nitrite ions bound in the active sites to NO species is observed, which is the beginning of the catalytic reaction. For the free form, an increase in the distance between the oxygen ligand bound to the catalytic heme and the iron ion of the heme takes place. In the sulfite complex no enzymatic reaction is detected
hanging drop vapour diffusion method
-
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
50 - 70
Q5F2I3
at least 50 h, purified enzyme, stable
80
Q5F2I3
the half-life is 2.2 h, enzyme inactivation is described by a two-step scheme, about 20% residual activity of after 25 h
86
Q5F2I3
melting of the TvNiR molecule is an irreversible highly cooperative process with the transition temperature of 86°C and the transition enthalpy of about J/mol of hexamer
30
-
at an enzyme concentration of 2 mg/ml in potassium phosphate buffer pH 7.0 the enzyme is stable at 30°C for at least 24 h
50
-
incubation at 50°C results in 1.5fold increase of activity
70
-
at 70°C the enzyme looses 50% of its activity in about 3 h
80
-
at 80°C activity decreases by 50% in 10min
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
30°C, potassium phosphate buffer at pH 7.0, 24 h, no loss of activity
-
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
native enzyme 57.7fold by anion exchange chromatography and gel filtration
Q5F2I3
non-denaturing preparative polyacrylamide gel electrophoresis
-
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Boyko, K.M.; Polyakov, K.M.; Tikhonova, T.V.; Slutsky, A.; Antipov, A.N.; Zvyagilskaya, R.A.; Bourenkov, G.P.; Popov, A.N.; Lamzin, V.S.; Popov, V.O.
Crystallization and preliminary X-ray analysis of cytochrome c nitrite reductase from Thioalkalivibrio nitratireducens
Acta Crystallogr. Sect. F
62
215-217
2006
Thioalkalivibrio nitratireducens, Thioalkalivibrio nitratireducens ALEN 2
Manually annotated by BRENDA team
Tikhonova, T.V.; Slutsky, A.; Antipov, A.N.; Boyko, K.M.; Polyakov, K.M.; Sorokin, D.Y.; Zvyagilskaya, R.A.; Popov, V.O.
Molecular and catalytic properties of a novel cytochrome c nitrite reductase from nitrate-reducing haloalkaliphilic sulfur-oxidizing bacterium Thioalkalivibrio nitratireducens
Biochim. Biophys. Acta
1764
715-723
2006
Thioalkalivibrio nitratireducens, Thioalkalivibrio nitratireducens ALEN 2
Manually annotated by BRENDA team
Tikhonova, T.V.; Slutskaya, E.S.; Filimonenkov, A.A.; Boyko, K.M.; Kleimenov, S.Y.; Konarev, P.V.; Polyakov, K.M.; Svergun, D.I.; Trofimov, A.A.; Khomenkov, V.G.; Zvyagilskaya, R.A.; Popov, V.O.
Isolation and oligomeric composition of cytochrome c nitrite reductase from the haloalkaliphilic bacterium Thioalkalivibrio nitratireducens
Biochemistry (Moscow)
73
164-170
2008
Thioalkalivibrio nitratireducens (Q5F2I3), Thioalkalivibrio nitratireducens
Manually annotated by BRENDA team
Polyakov, K.M.; Boyko, K.M.; Tikhonova, T.V.; Slutsky, A.; Antipov, A.N.; Zvyagilskaya, R.A.; Popov, A.N.; Bourenkov, G.P.; Lamzin, V.S.; Popov, V.O.
High-resolution structural analysis of a novel octaheme cytochrome c nitrite reductase from the haloalkaliphilic bacterium Thioalkalivibrio nitratireducens
J. Mol. Biol.
389
846-862
2009
Thioalkalivibrio nitratireducens (Q5F2I3), Thioalkalivibrio nitratireducens
Manually annotated by BRENDA team
Trofimov, A.A.; Polyakov, K.M.; Boyko, K.M.; Tikhonova, T.V.; Safonova, T.N.; Tikhonov, A.V.; Popov, A.N.; Popov, V.O.
Structures of complexes of octahaem cytochrome c nitrite reductase from Thioalkalivibrio nitratireducens with sulfite and cyanide
Acta Crystallogr. Sect. D
66
1043-1047
2010
Thioalkalivibrio nitratireducens (Q5F2I3), Thioalkalivibrio nitratireducens
Manually annotated by BRENDA team
Trofimov, A.A.; Polyakov, K.M.; Tikhonova, T.V.; Tikhonov, A.V.; Safonova, T.N.; Boyko, K.M.; Dorovatovskii, P.V.; Popov, V.O.
Covalent modifications of the catalytic tyrosine in octahaem cytochrome c nitrite reductase and their effect on the enzyme activity
Acta Crystallogr. Sect. D
68
144-153
2012
Thioalkalivibrio nitratireducens (Q5F2I3), Thioalkalivibrio nitratireducens
Manually annotated by BRENDA team
Trofimov, A.A.; Polyakov, K.M.; Lazarenko, V.A.; Popov, A.N.; Tikhonova, T.V.; Tikhonov, A.V.; Popov, V.O.
Structural study of the X-ray-induced enzymatic reaction of octahaem cytochrome C nitrite reductase
Acta Crystallogr. Sect. D
71
1087-1094
2015
Thioalkalivibrio nitratireducens (L0DSL2), Thioalkalivibrio nitratireducens, Thioalkalivibrio nitratireducens DSM 14787 (L0DSL2)
Manually annotated by BRENDA team