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Escherichia coli K-12 substr. MG1655 Enzyme: nitrite reductase

Synonyms: NirB

Subunit composition of nitrite reductase = [NirD][(NirB)2]
         nitrite reductase, small subunit = NirD (summary available)
         dimer of large subunit of nitrite reductase = (NirB)2
                 nitrite reductase, large subunit = NirB

Summary:
E. coli has two distinct nitrite reductases. Both enzymes reduce nitrite to ammonia. Their expresssion is complementary--with low concentrations of nitrate in the environment pentaheme nitrite reductase (NrfA) is made; with high concentrations of nitrate the NADH-dependent nitrite reductase (NirB) is made almost exclusively. At intermediate concentrations of nitrate, both are made. This regulation acts through the Nar regulatory circuit. Nitrite also induces the formation of both enzymes but it is a less effective inducer than nitrate by at least two orders of magnitude [Wang00].

NirB is located in the cytoplasm and does not generate a proton gradient. Its probable metabolic role is to detoxify nitrite. However, when cells are growing on high concentrations of nitrate (when NirB is active), nitrite nevertheless is excreted into the environment [Wang00].

NirB catalyzes the six-electron reduction of nitrite to ammonia and also catalyzes the two-electron reduction of hydroxylamine to ammonia. The prosthetic groups of nitrite reductase are FAD, an iron-sulfur cluster and siroheme [Jackson81, Macdonald85, Cammack82]. The product of the cysG gene is necessary for the synthesis of the siroheme prosthetic group of NirB. The reaction is active only during anaerobic growth and fulfills a dissimilatory rather than assimilatory role [Harborne92].

No NADH-dependent nitrite reductase activity was detected in E.coli strains lacking either NirB or NirD but a mixture of the two was as active as an extract from wild type [Harborne92]. Reconstitution of nitrite reductase activity in vitro required stoichiometric quantities of NirB and NirD, and NirD remained associated with NirB during the initial stages of enzyme purification suggesting that NirD is a structural subunit of the enzyme [Harborne92].

Locations: cytosol

Gene-Reaction Schematic: ?

GO Terms:

Biological Process: GO:0009061 - anaerobic respiration Inferred from experiment [Page90]
Molecular Function: GO:0008942 - nitrite reductase [NAD(P)H] activity Inferred from experiment Inferred by computational analysis [GOA01, Coleman78, Jackson81a]
Cellular Component: GO:0009344 - nitrite reductase complex [NAD(P)H] Inferred from experiment [Harborne92]
GO:0005737 - cytoplasm Inferred by computational analysis [GOA00]
GO:0005829 - cytosol

Credits:
Reviewed 17-Apr-2008 by Nolan L , Macquarie University
Last-Curated ? 30-Apr-2008 by Nolan L , Macquarie University


Enzymatic reaction of: nitrite reductase

Synonyms: nitrite reductase NADH, NADH:nitrite oxidoreductase

EC Number: 1.7.1.15

nitrite + 3 NADH + 5 H+ <=> ammonium + 3 NAD+ + 2 H2O

The reaction direction shown, that is, A + B ↔ C + D versus C + D ↔ A + B, is in accordance with the direction of enzyme catalysis.

The reaction is irreversible in the direction shown.

Alternative Substrates for nitrite: ferricyanide [Jackson81 ] , oxidized DCPIP [Jackson81 ] , hydroxylamine [Jackson81 ]

Cofactors or Prosthetic Groups: siroheme [Jackson81], FAD [Jackson81], [FeS] iron-sulfur cluster [Jackson81, Cammack82]

Inhibitors (Unknown Mechanism): NAD+ [Comment 1] , p-chloromercuribenzoate [Coleman78] , sulfite [Coleman78] , arsenite [Coleman78] , hydrogen cyanide [Coleman78]

Kinetic Parameters:

Substrate
Km (μM)
Citations
NADH
48.8
[Jackson82, BRENDA14]
nitrite
11.0
[Jackson82, BRENDA14]


Subunit of nitrite reductase: nitrite reductase, small subunit

Synonyms: NirD

Gene: nirD Accession Numbers: EG10655 (EcoCyc), b3366, ECK3354

Locations: cytosol

Sequence Length: 108 AAs

Molecular Weight: 12.284 kD (from nucleotide sequence)

pI: 5.17

GO Terms:

Biological Process: GO:0009061 - anaerobic respiration Inferred from experiment [Page90]
GO:0042128 - nitrate assimilation Inferred by computational analysis [UniProtGOA11a, GOA01a]
GO:0055114 - oxidation-reduction process Inferred by computational analysis [UniProtGOA11a, GOA01a]
Molecular Function: GO:0008942 - nitrite reductase [NAD(P)H] activity Inferred from experiment Inferred by computational analysis [GOA01a, Harborne92, Coleman78]
GO:0016491 - oxidoreductase activity Inferred by computational analysis [UniProtGOA11a, GOA01a]
GO:0051537 - 2 iron, 2 sulfur cluster binding Inferred by computational analysis [GOA01a]
Cellular Component: GO:0009344 - nitrite reductase complex [NAD(P)H] Inferred from experiment [Harborne92]
GO:0005737 - cytoplasm Inferred by computational analysis [UniProtGOA11, UniProtGOA11a]
GO:0005829 - cytosol Inferred by computational analysis [DiazMejia09]

MultiFun Terms: metabolism energy metabolism, carbon anaerobic respiration

Unification Links: EcoliWiki:b3366 , ModBase:P0A9I8 , PR:PRO_000023391 , Pride:P0A9I8 , Protein Model Portal:P0A9I8 , RefSeq:NP_417825 , SMR:P0A9I8 , String:511145.b3366 , UniProt:P0A9I8

Relationship Links: InterPro:IN-FAMILY:IPR012748 , InterPro:IN-FAMILY:IPR017881 , InterPro:IN-FAMILY:IPR017941 , PDB:Structure:2JO6 , Prosite:IN-FAMILY:PS51300

Summary:
The nirD gene encodes the small subunit of the nitrite reductase. [Peakman90, Harborne92]

Essentiality data for nirD knockouts: ?

Growth Medium Growth? T (°C) O2 pH Osm/L Growth Observations
LB enriched Yes 37 Aerobic 6.95   Yes [Gerdes03, Comment 2]
LB Lennox Yes 37 Aerobic 7   Yes [Baba06, Comment 3]
M9 medium with 1% glycerol Yes 37 Aerobic 7.2 0.35 Yes [Joyce06, Comment 4]
MOPS medium with 0.4% glucose Yes 37 Aerobic 7.2 0.22 Yes [Baba06, Comment 3]
Yes [Feist07, Comment 5]

Subunit of nitrite reductase: dimer of large subunit of nitrite reductase

Gene: nirB Accession Numbers: EG10653 (EcoCyc), b3365, ECK3353

Locations: cytosol

Subunit composition of dimer of large subunit of nitrite reductase = [NirB]2
         nitrite reductase, large subunit = NirB

Map Position: [3,492,033 -> 3,494,576] (75.26 centisomes)
Length: 2544 bp / 847 aa

Molecular Weight of Polypeptide: 93.121 kD (from nucleotide sequence)

pI: 6.23

GO Terms:

Biological Process: GO:0009061 - anaerobic respiration Inferred from experiment [Page90]
GO:0042128 - nitrate assimilation Inferred by computational analysis [UniProtGOA12, UniProtGOA11a, GOA01a]
GO:0055114 - oxidation-reduction process Inferred by computational analysis [UniProtGOA11a, GOA01a]
Molecular Function: GO:0008942 - nitrite reductase [NAD(P)H] activity Inferred from experiment Inferred by computational analysis [GOA01a, Harborne92, Coleman78]
GO:0020037 - heme binding Inferred from experiment Inferred by computational analysis [GOA01a, Jackson81]
GO:0050660 - flavin adenine dinucleotide binding Inferred from experiment Inferred by computational analysis [GOA01a, Jackson81]
GO:0050661 - NADP binding Inferred from experiment Inferred by computational analysis [GOA01a, Coleman78]
GO:0051536 - iron-sulfur cluster binding Inferred from experiment Inferred by computational analysis [UniProtGOA11a, GOA01a, Cammack82]
GO:0016491 - oxidoreductase activity Inferred by computational analysis [UniProtGOA11a, GOA01a]
GO:0046872 - metal ion binding Inferred by computational analysis [UniProtGOA11a]
GO:0051539 - 4 iron, 4 sulfur cluster binding Inferred by computational analysis [UniProtGOA11a]
Cellular Component: GO:0009344 - nitrite reductase complex [NAD(P)H] Inferred from experiment [Coleman78, Jackson81a]
GO:0005737 - cytoplasm Inferred by computational analysis [GOA00]
GO:0005829 - cytosol Inferred by computational analysis [DiazMejia09, Page90]

MultiFun Terms: metabolism energy metabolism, carbon anaerobic respiration

Unification Links: EcoliWiki:b3365 , ModBase:P08201 , Protein Model Portal:P08201 , RefSeq:NP_417824 , SMR:P08201 , String:511145.b3365 , UniProt:P08201

Relationship Links: InterPro:IN-FAMILY:IPR001327 , InterPro:IN-FAMILY:IPR005117 , InterPro:IN-FAMILY:IPR006066 , InterPro:IN-FAMILY:IPR006067 , InterPro:IN-FAMILY:IPR007419 , InterPro:IN-FAMILY:IPR012744 , InterPro:IN-FAMILY:IPR013027 , InterPro:IN-FAMILY:IPR017121 , InterPro:IN-FAMILY:IPR023753 , Pfam:IN-FAMILY:PF00070 , Pfam:IN-FAMILY:PF01077 , Pfam:IN-FAMILY:PF03460 , Pfam:IN-FAMILY:PF04324 , Pfam:IN-FAMILY:PF07992 , Prints:IN-FAMILY:PR00368 , Prints:IN-FAMILY:PR00397 , Prosite:IN-FAMILY:PS00365

Summary:
Deletion of the nirB gene increases the response to nitrite, which decreases NO accumulation in the cytoplasm, protecting bacteria against nitrosative stress [Vine11].

Gene Citations: [Peakman90a, Harborne92]

Essentiality data for nirB knockouts: ?

Growth Medium Growth? T (°C) O2 pH Osm/L Growth Observations
LB enriched Yes 37 Aerobic 6.95   Yes [Gerdes03, Comment 2]
LB Lennox Yes 37 Aerobic 7   Yes [Baba06, Comment 3]
M9 medium with 1% glycerol Yes 37 Aerobic 7.2 0.35 Yes [Joyce06, Comment 4]
MOPS medium with 0.4% glucose Yes 37 Aerobic 7.2 0.22 Yes [Baba06, Comment 3]
Yes [Feist07, Comment 5]

References

Baba06: Baba T, Ara T, Hasegawa M, Takai Y, Okumura Y, Baba M, Datsenko KA, Tomita M, Wanner BL, Mori H (2006). "Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection." Mol Syst Biol 2;2006.0008. PMID: 16738554

BRENDA14: BRENDA team (2014). "Imported from BRENDA version existing on Aug 2014." http://www.brenda-enzymes.org.

Cammack82: Cammack R, Jackson RH, Cornish-Bowden A, Cole JA (1982). "Electron-spin-resonance studies of the NADH-dependent nitrite reductase from Escherichia coli K12." Biochem J 207(2);333-9. PMID: 6297458

Coleman78: Coleman KJ, Cornish-Bowden A, Cole JA (1978). "Purification and properties of nitrite reductase from Escherichia coli K12." Biochem J 1978;175(2);483-93. PMID: 217342

DiazMejia09: Diaz-Mejia JJ, Babu M, Emili A (2009). "Computational and experimental approaches to chart the Escherichia coli cell-envelope-associated proteome and interactome." FEMS Microbiol Rev 33(1);66-97. PMID: 19054114

Feist07: Feist AM, Henry CS, Reed JL, Krummenacker M, Joyce AR, Karp PD, Broadbelt LJ, Hatzimanikatis V, Palsson BO (2007). "A genome-scale metabolic reconstruction for Escherichia coli K-12 MG1655 that accounts for 1260 ORFs and thermodynamic information." Mol Syst Biol 3;121. PMID: 17593909

Gerdes03: Gerdes SY, Scholle MD, Campbell JW, Balazsi G, Ravasz E, Daugherty MD, Somera AL, Kyrpides NC, Anderson I, Gelfand MS, Bhattacharya A, Kapatral V, D'Souza M, Baev MV, Grechkin Y, Mseeh F, Fonstein MY, Overbeek R, Barabasi AL, Oltvai ZN, Osterman AL (2003). "Experimental determination and system level analysis of essential genes in Escherichia coli MG1655." J Bacteriol 185(19);5673-84. PMID: 13129938

GOA00: GOA (2000). "Gene Ontology annotation based on Swiss-Prot keyword mapping."

GOA01: GOA, MGI (2001). "Gene Ontology annotation based on Enzyme Commission mapping." Genomics 74;121-128.

GOA01a: GOA, DDB, FB, MGI, ZFIN (2001). "Gene Ontology annotation through association of InterPro records with GO terms."

Harborne92: Harborne NR, Griffiths L, Busby SJ, Cole JA (1992). "Transcriptional control, translation and function of the products of the five open reading frames of the Escherichia coli nir operon." Mol Microbiol 1992;6(19);2805-13. PMID: 1435259

Jackson81: Jackson RH, Cornish-Bowden A, Cole JA (1981). "Prosthetic groups of the NADH-dependent nitrite reductase from Escherichia coli K12." Biochem J 1981;193(3);861-7. PMID: 7030314

Jackson81a: Jackson RH, Cole JA, Cornish-Bowden A (1981). "The steady-state kinetics of the NADH-dependent nitrite reductase from Escherichia coli K 12. Nitrite and hydroxylamine reduction." Biochem J 1981;199(1);171-8. PMID: 6279095

Jackson82: Jackson RH, Cole JA, Cornish-Bowden A (1982). "The steady state kinetics of the NADH-dependent nitrite reductase from Escherichia coli K12. The reduction of single-electron acceptors." Biochem J 203(2);505-10. PMID: 6288003

Joyce06: Joyce AR, Reed JL, White A, Edwards R, Osterman A, Baba T, Mori H, Lesely SA, Palsson BO, Agarwalla S (2006). "Experimental and computational assessment of conditionally essential genes in Escherichia coli." J Bacteriol 188(23);8259-71. PMID: 17012394

Macdonald85: Macdonald H, Cole J (1985). "Molecular cloning and functional analysis of the cysG and nirB genes of Escherichia coli K12, two closely-linked genes required for NADH-dependent nitrite reductase activity." Mol Gen Genet 1985;200(2);328-34. PMID: 2993824

Page90: Page L, Griffiths L, Cole JA (1990). "Different physiological roles of two independent pathways for nitrite reduction to ammonia by enteric bacteria." Arch Microbiol 1990;154(4);349-54. PMID: 2173895

Peakman90: Peakman T, Crouzet J, Mayaux JF, Busby S, Mohan S, Harborne N, Wootton J, Nicolson R, Cole J (1990). "Nucleotide sequence, organisation and structural analysis of the products of genes in the nirB-cysG region of the Escherichia coli K-12 chromosome." Eur J Biochem 1990;191(2);315-23. PMID: 2200672

Peakman90a: Peakman T, Busby S, Cole J (1990). "Transcriptional control of the cysG gene of Escherichia coli K-12 during aerobic and anaerobic growth." Eur J Biochem 1990;191(2);325-31. PMID: 2200673

UniProtGOA11: UniProt-GOA (2011). "Gene Ontology annotation based on the manual assignment of UniProtKB Subcellular Location terms in UniProtKB/Swiss-Prot entries."

UniProtGOA11a: UniProt-GOA (2011). "Gene Ontology annotation based on manual assignment of UniProtKB keywords in UniProtKB/Swiss-Prot entries."

UniProtGOA12: UniProt-GOA (2012). "Gene Ontology annotation based on UniPathway vocabulary mapping."

Vine11: Vine CE, Purewal SK, Cole JA (2011). "NsrR-dependent method for detecting nitric oxide accumulation in the Escherichia coli cytoplasm and enzymes involved in NO production." FEMS Microbiol Lett 325(2);108-14. PMID: 22092912

Wang00: Wang H, Gunsalus RP (2000). "The nrfA and nirB nitrite reductase operons in Escherichia coli are expressed differently in response to nitrate than to nitrite." J Bacteriol 182(20);5813-22. PMID: 11004182


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Please cite the following article in publications resulting from the use of EcoCyc: Nucleic Acids Research 41:D605-12 2013
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