Escherichia coli K-12 substr. MG1655 Polypeptide: large subunit of periplasmic nitrate reductase, molybdoprotein

Gene: napA Accession Numbers: EG12067 (EcoCyc), b2206, ECK2198

Synonyms: yojD, yojC, yojE

Regulation Summary Diagram

Regulation summary diagram for napA

Component of: periplasmic nitrate reductase (extended summary available)

The napA gene encodes the periplasmic nitrate reductase molybdoprotein with an Fe-S center [Grove96, Cole96]. NapA binds a [4Fe-4S] cluster and bis-molybdopterin guanine dinucleotide [Jepson07]. NapA is synthesized as a precursor with a 36 residue signal peptide that is removed upon export to the periplasm [Thomas99]. Prior to export the the signal peptide of NapA is bound to the NapD chaperone which may act to protect NapD from translocation until cofactor loading is complete [Maillard07]. A NapD binding epitope is located towards the N-terminus of the NapA signal peptide and partially overlaps with the twin arginine motif [Grahl12]. An NMR structure of the NapD-NapA signal peptide complex has been reported [Grahl12].

Gene Citations: [Darwin95]

Locations: periplasmic space

Map Position: [2,300,267 <- 2,302,753] (49.56 centisomes, 178°)
Length: 2487 bp / 828 aa

Molecular Weight of Polypeptide: 93.042 kD (from nucleotide sequence)

Unification Links: ASAP:ABE-0007289, CGSC:36550, DIP:DIP-10304N, EchoBASE:EB1994, EcoGene:EG12067, EcoliWiki:b2206, Mint:MINT-1243254, ModBase:P33937, OU-Microarray:b2206, PortEco:napA, PR:PRO_000023345, Pride:P33937, Protein Model Portal:P33937, RefSeq:NP_416710, RegulonDB:EG12067, SMR:P33937, String:511145.b2206, UniProt:P33937

Relationship Links: InterPro:IN-FAMILY:IPR006311, InterPro:IN-FAMILY:IPR006656, InterPro:IN-FAMILY:IPR006657, InterPro:IN-FAMILY:IPR006963, InterPro:IN-FAMILY:IPR009010, InterPro:IN-FAMILY:IPR010051, InterPro:IN-FAMILY:IPR019546, InterPro:IN-FAMILY:IPR027467, PDB:Structure:2NYA, PDB:Structure:2PQ4, Pfam:IN-FAMILY:PF00384, Pfam:IN-FAMILY:PF01568, Pfam:IN-FAMILY:PF04879, Prosite:IN-FAMILY:PS00551, Prosite:IN-FAMILY:PS51318, Prosite:IN-FAMILY:PS51669, Smart:IN-FAMILY:SM00926

In Paralogous Gene Group: 222 (14 members)

Gene-Reaction Schematic

Gene-Reaction Schematic

Genetic Regulation Schematic

Genetic regulation schematic for napA

GO Terms:
Biological Process:
Inferred from experimentGO:0009061 - anaerobic respiration [Stewart02]
Inferred by computational analysisGO:0006777 - Mo-molybdopterin cofactor biosynthetic process [GOA06]
Inferred by computational analysisGO:0042128 - nitrate assimilation [UniProtGOA11a, GOA06, GOA01a]
Inferred by computational analysisGO:0055114 - oxidation-reduction process [UniProtGOA11a, GOA06, GOA01a]
Molecular Function:
Inferred from experimentGO:0005515 - protein binding [Rajagopala14, Dow14, Chan10a, Chan09]
Inferred from experimentInferred by computational analysisGO:0008940 - nitrate reductase activity [GOA06, GOA01, GOA01a, Thomas99, Stewart02, Grove96]
Inferred from experimentInferred by computational analysisGO:0030151 - molybdenum ion binding [GOA01a, Jepson07]
Inferred from experimentInferred by computational analysisGO:0051539 - 4 iron, 4 sulfur cluster binding [UniProtGOA11a, GOA06, GOA01a, Jepson07]
Inferred by computational analysisGO:0003954 - NADH dehydrogenase activity [Gaudet10]
Inferred by computational analysisGO:0005506 - iron ion binding [GOA06]
Inferred by computational analysisGO:0009055 - electron carrier activity [GOA06]
Inferred by computational analysisGO:0016491 - oxidoreductase activity [UniProtGOA11a, GOA01a]
Inferred by computational analysisGO:0046872 - metal ion binding [UniProtGOA11a]
Inferred by computational analysisGO:0051536 - iron-sulfur cluster binding [UniProtGOA11a]
Cellular Component:
Inferred from experimentInferred by computational analysisGO:0030288 - outer membrane-bounded periplasmic space [DiazMejia09, Han14, Thomas99]
Inferred from experimentInferred by computational analysisGO:0042597 - periplasmic space [UniProtGOA11, UniProtGOA11a, GOA06, GOA01a, Grove96]

MultiFun Terms: metabolismenergy metabolism, carbonanaerobic respiration
metabolismenergy production/transportelectron acceptors

Essentiality data for napA knockouts:

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

Last-Curated 06-Mar-2012 by Mackie A, Macquarie University

Subunit of: periplasmic nitrate reductase

Synonyms: Nap

Subunit composition of periplasmic nitrate reductase = [NapB][NapC][(NapG)(NapH)][NapA]
         subunit of periplasmic nitrate reductase, cytochrome c550 protein = NapB (summary available)
         periplasmic nitrate reductase, cytochrome c protein = NapC (summary available)
         NapGH, alternative quinol dehydrogenase = (NapG)(NapH) (summary available)
                 ferredoxin-type protein = NapG (summary available)
                 ferredoxin-type protein = NapH (summary available)
         large subunit of periplasmic nitrate reductase, molybdoprotein = NapA (summary available)

E. coli K-12 contains three nitrate reductases. Two of them, nitrate reductase A (NRA) and nitrate reductase Z (NRZ), are membrane bound and biochemically similar. The third nitrate reductase, Nap, is located in the periplasm. It is induced by anaerobiosis through the mediation of the transcription factor Fnr and low concentrations of nitrate through the mediation of NarP [McNicholas02]. Nap is not itself a coupling site for generating proton motive force; acting as a terminal electron acceptor, it does support anaerobic respiration of various carbon sources [Stewart02].

The physiological role of Nap is that of mediating anaerobic respiration at the expense of low concentrations of nitrate. Owing to the periplasmic location of Nap, the cost of pumping nitrate into the cell is avoided. In addition, Nap has a significantly higher affinity for nitrate than NRA and is thus able to exploit the low concentrations of nitrate occuring in the natural environment of E. coli [Potter99]. Notably, several pathogenic bacterial species, such as Haemophilus influenzae, only contain orthologs of the periplasmic nitrate reductase [Potter99]. During glucose fermentation in the absence of menaquinone, a very low level of Nap activity appears to substitute for the redox-balancing role of fumarate reductase, which is dependent on menaquinone [Brondijk04].

The nap operon encodes seven proteins. The catalytic portion of the protein, consisting of the periplasmic NapA and NapB polypeptides, receives electrons via the membrane-bound cytochrome NapC from NapGH or directly from the quinone pool. The NapD polypeptide is required for enzyme activity and is thought to be involved in the post-translational assembly of the molybdoprotein NapA. NapF, NapG and NapH are predicted to encode iron-sulfur proteins and are not required for Nap activity; they do, however, contribute to the maximum rate of nitrate reduction. NapG and NapH facilitate electron transfer from ubiquinol via NapC to NapAB. [Brondijk04, Brondijk02, Potter99a, Cole96, Grove96, Grove96a, IobbiNivol94]

nap: nitrate reductase in the periplasm [Grove96]

Citations: [Dow14]

Locations: periplasmic space

GO Terms:
Cellular Component:
GO:0030288 - outer membrane-bounded periplasmic space []

Enzymatic reaction of: ubiquinol:nitrate oxidoreductase (periplasmic) (periplasmic nitrate reductase)

Inferred from experiment

EC Number:

Transport reaction diagram for ubiquinol:nitrate oxidoreductase (periplasmic)

In Pathways: nitrate reduction X (periplasmic, dissimilatory)

Cofactors or Prosthetic Groups: heme c [Jepson07], a [4Fe-4S] iron-sulfur cluster [Jepson07], bis(guanylyl molybdopterin cofactor) [Jepson07]

Enzymatic reaction of: menaquinol:nitrate oxidoreductase (periplasmic) (periplasmic nitrate reductase)

Inferred from experiment

EC Number:

Transport reaction diagram for menaquinol:nitrate oxidoreductase (periplasmic)

Sequence Features

Protein sequence of large subunit of periplasmic nitrate reductase, molybdoprotein with features indicated

Feature Class Location Citations Comment
Signal-Sequence 1 -> 36
Inferred from experiment[Thomas99]
UniProt: Tat-type signal.
Chain 37 -> 828
Author statement[UniProt15]
UniProt: Periplasmic nitrate reductase.
Conserved-Region 39 -> 95
Author statement[UniProt15]
UniProt: 4Fe-4S Mo/W bis-MGD-type.
Metal-Binding-Site 46
Inferred by computational analysis[UniProt15]
UniProt: Iron-sulfur (4Fe-4S).
Metal-Binding-Site 49
Inferred by computational analysis[UniProt15]
UniProt: Iron-sulfur (4Fe-4S).
Metal-Binding-Site 53
Inferred by computational analysis[UniProt15]
UniProt: Iron-sulfur (4Fe-4S).
Metal-Binding-Site 81
Inferred by computational analysis[UniProt15]
UniProt: Iron-sulfur (4Fe-4S).
Sequence-Conflict 98
Inferred by curator[UniProt15]
UniProt: (in Ref. 1; AAA16399).

Sequence Pfam Features

Protein sequence of large subunit of periplasmic nitrate reductase, molybdoprotein with features indicated

Feature Class Location Citations Comment
Pfam PF04879 40 -> 92
Inferred by computational analysis[Finn14]
Molybdop_Fe4S4 : Molybdopterin oxidoreductase Fe4S4 domain [More...]
Pfam PF00384 96 -> 567
Inferred by computational analysis[Finn14]
Molybdopterin : Molybdopterin oxidoreductase
Pfam PF01568 714 -> 822
Inferred by computational analysis[Finn14]
Molydop_binding : Molydopterin dinucleotide binding domain [More...]

Gene Local Context (not to scale -- see Genome Browser for correct scale)

Transcription Units

Transcription-unit diagram

Transcription-unit diagram

Transcription-unit diagram

Transcription-unit diagram


10/20/97 Gene b2206 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG12067; confirmed by SwissProt match.


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

Brondijk02: Brondijk TH, Fiegen D, Richardson DJ, Cole JA (2002). "Roles of NapF, NapG and NapH, subunits of the Escherichia coli periplasmic nitrate reductase, in ubiquinol oxidation." Mol Microbiol 44(1);245-55. PMID: 11967083

Brondijk04: Brondijk TH, Nilavongse A, Filenko N, Richardson DJ, Cole JA (2004). "NapGH components of the periplasmic nitrate reductase of Escherichia coli K-12: location, topology and physiological roles in quinol oxidation and redox balancing." Biochem J 379(Pt 1);47-55. PMID: 14674886

Chan09: Chan CS, Chang L, Rommens KL, Turner RJ (2009). "Differential interactions between Tat-specific redox enzyme peptides and their chaperones." J Bacteriol 191(7):2091-101. PMID: 19151138

Chan10a: Chan CS, Chang L, Winstone TM, Turner RJ (2010). "Comparing system-specific chaperone interactions with their Tat dependent redox enzyme substrates." FEBS Lett 584(22);4553-8. PMID: 20974141

Cole96: Cole J (1996). "Nitrate reduction to ammonia by enteric bacteria: redundancy, or a strategy for survival during oxygen starvation?." FEMS Microbiol Lett 1996;136(1);1-11. PMID: 8919448

Darwin95: Darwin AJ, Stewart V (1995). "Nitrate and nitrite regulation of the Fnr-dependent aeg-46.5 promoter of Escherichia coli K-12 is mediated by competition between homologous response regulators (NarL and NarP) for a common DNA-binding site." J Mol Biol 1995;251(1);15-29. PMID: 7643383

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

Dow14: Dow JM, Grahl S, Ward R, Evans R, Byron O, Norman DG, Palmer T, Sargent F (2014). "Characterization of a periplasmic nitrate reductase in complex with its biosynthetic chaperone." FEBS J 281(1);246-60. PMID: 24314029

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

Finn14: Finn RD, Bateman A, Clements J, Coggill P, Eberhardt RY, Eddy SR, Heger A, Hetherington K, Holm L, Mistry J, Sonnhammer EL, Tate J, Punta M (2014). "Pfam: the protein families database." Nucleic Acids Res 42(Database issue);D222-30. PMID: 24288371

Gaudet10: Gaudet P, Livstone M, Thomas P (2010). "Annotation inferences using phylogenetic trees." PMID: 19578431

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

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."

GOA06: GOA, SIB (2006). "Electronic Gene Ontology annotations created by transferring manual GO annotations between orthologous microbial proteins."

Grahl12: Grahl S, Maillard J, Spronk CA, Vuister GW, Sargent F (2012). "Overlapping transport and chaperone-binding functions within a bacterial twin-arginine signal peptide." Mol Microbiol 83(6);1254-67. PMID: 22329966

Grove96: Grove J, Tanapongpipat S, Thomas G, Griffiths L, Crooke H, Cole J (1996). "Escherichia coli K-12 genes essential for the synthesis of c-type cytochromes and a third nitrate reductase located in the periplasm." Mol Microbiol 1996;19(3);467-81. PMID: 8830238

Grove96a: Grove J, Busby S, Cole J (1996). "The role of the genes nrf EFG and ccmFH in cytochrome c biosynthesis in Escherichia coli." Mol Gen Genet 1996;252(3);332-41. PMID: 8842153

Han14: Han MJ, Kim JY, Kim JA (2014). "Comparison of the large-scale periplasmic proteomes of the Escherichia coli K-12 and B strains." J Biosci Bioeng 117(4);437-42. PMID: 24140104

IobbiNivol94: Iobbi-Nivol C, Crooke H, Griffiths L, Grove J, Hussain H, Pommier J, Mejean V, Cole JA (1994). "A reassessment of the range of c-type cytochromes synthesized by Escherichia coli K-12." FEMS Microbiol Lett 1994;119(1-2);89-94. PMID: 8039676

Jepson07: Jepson BJ, Mohan S, Clarke TA, Gates AJ, Cole JA, Butler CS, Butt JN, Hemmings AM, Richardson DJ (2007). "Spectropotentiometric and structural analysis of the periplasmic nitrate reductase from Escherichia coli." J Biol Chem 282(9);6425-37. PMID: 17130127

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

Maillard07: Maillard J, Spronk CA, Buchanan G, Lyall V, Richardson DJ, Palmer T, Vuister GW, Sargent F (2007). "Structural diversity in twin-arginine signal peptide-binding proteins." Proc Natl Acad Sci U S A 104(40);15641-6. PMID: 17901208

McNicholas02: McNicholas PM, Gunsalus RP (2002). "The molybdate-responsive Escherichia coli ModE transcriptional regulator coordinates periplasmic nitrate reductase (napFDAGHBC) operon expression with nitrate and molybdate availability." J Bacteriol 184(12);3253-9. PMID: 12029041

Park06: Park YJ, Yoo CB, Choi SY, Lee HB (2006). "Purifications and characterizations of a ferredoxin and its related 2-oxoacid:ferredoxin oxidoreductase from the hyperthermophilic archaeon, Sulfolobus solfataricus P1." J Biochem Mol Biol 39(1);46-54. PMID: 16466637

Potter99: Potter LC, Millington P, Griffiths L, Thomas GH, Cole JA (1999). "Competition between Escherichia coli strains expressing either a periplasmic or a membrane-bound nitrate reductase: does Nap confer a selective advantage during nitrate-limited growth?." Biochem J 344 Pt 1;77-84. PMID: 10548536

Potter99a: Potter LC, Cole JA (1999). "Essential roles for the products of the napABCD genes, but not napFGH, in periplasmic nitrate reduction by Escherichia coli K-12." Biochem J 1999;344 Pt 1;69-76. PMID: 10548535

Rajagopala14: Rajagopala SV, Sikorski P, Kumar A, Mosca R, Vlasblom J, Arnold R, Franca-Koh J, Pakala SB, Phanse S, Ceol A, Hauser R, Siszler G, Wuchty S, Emili A, Babu M, Aloy P, Pieper R, Uetz P (2014). "The binary protein-protein interaction landscape of Escherichia coli." Nat Biotechnol 32(3);285-90. PMID: 24561554

Stewart02: Stewart V, Lu Y, Darwin AJ (2002). "Periplasmic nitrate reductase (NapABC enzyme) supports anaerobic respiration by Escherichia coli K-12." J Bacteriol 184(5);1314-23. PMID: 11844760

Thomas99: Thomas G, Potter L, Cole JA (1999). "The periplasmic nitrate reductase from Escherichia coli: a heterodimeric molybdoprotein with a double-arginine signal sequence and an unusual leader peptide cleavage site." FEMS Microbiol Lett 1999;174(1);167-71. PMID: 10234835

UniProt15: UniProt Consortium (2015). "UniProt version 2015-08 released on 2015-07-22." Database.

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."

Other References Related to Gene Regulation

Choe93: Choe M, Reznikoff WS (1993). "Identification of the regulatory sequence of anaerobically expressed locus aeg-46.5." J Bacteriol 1993;175(4);1165-72. PMID: 8432709

Darwin98: Darwin AJ, Ziegelhoffer EC, Kiley PJ, Stewart V (1998). "Fnr, NarP, and NarL regulation of Escherichia coli K-12 napF (periplasmic nitrate reductase) operon transcription in vitro." J Bacteriol 1998;180(16);4192-8. PMID: 9696769

Giel06: Giel JL, Rodionov D, Liu M, Blattner FR, Kiley PJ (2006). "IscR-dependent gene expression links iron-sulphur cluster assembly to the control of O-regulated genes in Escherichia coli." Mol Microbiol 60(4);1058-75. PMID: 16677314

Partridge09: Partridge JD, Bodenmiller DM, Humphrys MS, Spiro S (2009). "NsrR targets in the Escherichia coli genome: new insights into DNA sequence requirements for binding and a role for NsrR in the regulation of motility." Mol Microbiol 73(4);680-94. PMID: 19656291

Pruss01: Pruss BM, Liu X, Hendrickson W, Matsumura P (2001). "FlhD/FlhC-regulated promoters analyzed by gene array and lacZ gene fusions." FEMS Microbiol Lett 2001;197(1);91-7. PMID: 11287152

Stewart03: Stewart V, Bledsoe PJ, Williams SB (2003). "Dual overlapping promoters control napF (periplasmic nitrate reductase) operon expression in Escherichia coli K-12." J Bacteriol 185(19);5862-70. PMID: 13129959

Stewart03a: Stewart V, Bledsoe PJ (2003). "Synthetic lac operator substitutions for studying the nitrate- and nitrite-responsive NarX-NarL and NarQ-NarP two-component regulatory systems of Escherichia coli K-12." J Bacteriol 185(7);2104-11. PMID: 12644479

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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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