|Gene:||wrbA||Accession Numbers: EG11540 (MetaCyc), b1004, ECK0995|
Species: Escherichia coli K-12 substr. MG1655
The purified WrbA protein has NAD(P)H:quinone oxidoreductase activity [Patridge06]. WrbA is related to the flavodoxin family of proteins [Grandori94]. Unlike the flavodoxins, WrbA does not have a stabilized semiquinone state. It rapidly takes up two electrons, generating the fully reduced form [Noll06]. Purified WrbA protein binds one FMN per monomer with a binding constant of 2 µM at room temperature, which is weaker than that of typical flavodoxins [Grandori98, Patridge06]. Binding of FMN appears to be pH-dependent [Patridge06], and it increases the thermal stability and promotes tetramerization of WrbA [Natalello07].
WrbA is a multimer in solution, existing in an equilibrium between the dimeric and tetrameric form [Grandori98, Patridge06]. Crystal structures show WrbA to be a dimer of dimers. Structural comparisons to flavodoxin and the mammalian NAD(P)H:quinone oxidoreductase Nqo1 allow interpretation of the differences in the cofactor requirements and the catalytic functions of these proteins [Wolfova07, Carey07]. Additional crystal structures of WrbA in complex with benzoquinone or NADH suggest that binding of quinones and NADH to the FMN cofactor is mutually exclusive [Andrade07]. Steady-state kinetic analysis suggests a ping-pong reaction mechanism and show two-plateau Michaelis-Menten plots that are dependent on the temperature at which the enzyme had been held. This result implies allosteric regulation of the enzyme [Kishko12].
Comparison of the crystal structures of the apo- and holoenzyme forms of WrbA led to improved understanding of the functional similarities and differences of WrbA compared to the flavodoxins [Wolfova09].
It was initially reported that WrbA copurifies with the Trp repressor protein TrpR and enhances the formation or stability of TrpR binding to its operator target sites [Yang93a]. However, a later report showed that WrbA does not specifically affect the DNA binding affinity or mode of binding of TrpR [Grandori98]. WrbA alone does not bind to the trp operator DNA [Yang93a]. The association between WrbA and TrpR observed by [Yang93a] may therefore be due to structural rather than functional reasons [Grandori98].
Expression of wrbA is increased during stationary phase and is RpoS-dependent [Yang93a, Lacour & Landini, 2004] and Crl-dependent [Lelong et al., 2007]. In a strain lacking the ClpP serine protease, the level of WrbA protein is decreased during exponential growth and late stationary phase [Weichart03]. Like other members of the RpoS regulon, the steady-state level of WrbA is increased by growth on acetate [Kirkpatrick01]. wrbA was predicted to be a target of the small RNA OxyS, and overexpression of OxyS decreases the expression of wrbA [Tjaden06]. Expression of wrbA is also likely directly repressed by ArcA-P [Liu04a].
A wrbA null mutant has no growth defect when assayed with the Biolog system. Growth of the mutant strain is inhibited by N-trichloromethyl-mercapto-4-cyclohexene-1,2-dicarboximide and 8-hydroxyquinoline [Patridge06].
WrbA: "tryptophan (W) repressor-binding protein" [Yang93a]
Locations: cytosol, membrane
|Map Position: [1,066,335 <- 1,066,931]|
Molecular Weight of Polypeptide: 20.846 kD (from nucleotide sequence), 21 kD (experimental) [Yang93a ]
Unification Links: ASAP:ABE-0003392 , CGSC:31836 , DIP:DIP-36231N , EchoBASE:EB1502 , EcoGene:EG11540 , EcoliWiki:b1004 , Mint:MINT-1272067 , ModBase:P0A8G6 , OU-Microarray:b1004 , PortEco:wrbA , PR:PRO_000024221 , Pride:P0A8G6 , Protein Model Portal:P0A8G6 , RefSeq:NP_415524 , RegulonDB:EG11540 , SMR:P0A8G6 , String:511145.b1004 , UniProt:P0A8G6
Relationship Links: InterPro:IN-FAMILY:IPR008254 , InterPro:IN-FAMILY:IPR010089 , InterPro:IN-FAMILY:IPR029039 , PDB:Structure:2R96 , PDB:Structure:2R97 , PDB:Structure:2RG1 , PDB:Structure:3B6I , PDB:Structure:3B6J , PDB:Structure:3B6K , PDB:Structure:3B6M , PDB:Structure:3ZHO , Pfam:IN-FAMILY:PF00258 , Prosite:IN-FAMILY:PS50902
Instance reactions of [an electron-transfer quinone[inner membrane] + NAD(P)H + H+ → an electron-transfer quinol[inner membrane] + NAD(P)+] (18.104.22.168):
|Biological Process:||GO:0006979 - response to oxidative stress
[Lacour & Landini, 2004]
GO:0045892 - negative regulation of transcription, DNA-templated [GOA01a]
GO:0055114 - oxidation-reduction process [UniProtGOA11a]
|Molecular Function:||GO:0003955 - NAD(P)H dehydrogenase (quinone) activity
[GOA06, GOA01, Patridge06]
GO:0010181 - FMN binding [GOA01a, Patridge06]
GO:0042802 - identical protein binding [Lasserre06]
GO:0000166 - nucleotide binding [UniProtGOA11a]
GO:0016491 - oxidoreductase activity [UniProtGOA11a, GOA01a]
GO:0050660 - flavin adenine dinucleotide binding [GOA06]
GO:0050661 - NADP binding [GOA06]
GO:0051287 - NAD binding [GOA06]
|Cellular Component:||GO:0005829 - cytosol
[Ishihama08, LopezCampistrou05, Patridge06]
GO:0016020 - membrane [Lasserre06]
GO:0005737 - cytoplasm
Enzymatic reaction of: NAD(P)H:quinone oxidoreductase
EC Number: 22.214.171.124
Alternative Substrates for an electron-transfer quinone: menadione [Patridge06 ] , 2,3-dihydroxy-5-methyl-1,4-benzoquinone [Patridge06 ] , 1,4-naphthoquinone [Patridge06 ] , 1,4-benzoquinone [Patridge06 ]
NADH is the preferred electron donor. Km values were measured for NADH and p-benzoquinone [Patridge06].
The midpoint potential at pH 7 is -115 mV [Zafar09].
pH(opt): 6-8 [Patridge06]
|Feature Class||Location||Attached Group||Citations||Comment|
|Chain||2 -> 198|
|Conserved-Region||4 -> 189|
|Nucleotide-Phosphate-Binding-Region||9 -> 14||FMN|
|Nucleotide-Phosphate-Binding-Region||77 -> 80||FMN|
|Nucleotide-Phosphate-Binding-Region||112 -> 118||FMN|
10/20/97 Gene b1004 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG11540; confirmed by SwissProt match.
Carey07: Carey J, Brynda J, Wolfova J, Grandori R, Gustavsson T, Ettrich R, Smatanova IK (2007). "WrbA bridges bacterial flavodoxins and eukaryotic NAD(P)H:quinone oxidoreductases." Protein Sci 16(10);2301-5. PMID: 17893367
Grandori98: Grandori R, Khalifah P, Boice JA, Fairman R, Giovanielli K, Carey J (1998). "Biochemical characterization of WrbA, founding member of a new family of multimeric flavodoxin-like proteins." J Biol Chem 273(33);20960-6. PMID: 9694845
Kirkpatrick01: Kirkpatrick C, Maurer LM, Oyelakin NE, Yoncheva YN, Maurer R, Slonczewski JL (2001). "Acetate and formate stress: opposite responses in the proteome of Escherichia coli." J Bacteriol 183(21);6466-77. PMID: 11591692
Kishko12: Kishko I, Harish B, Zayats V, Reha D, Tenner B, Beri D, Gustavsson T, Ettrich R, Carey J (2012). "Biphasic kinetic behavior of E. coli WrbA, an FMN-dependent NAD(P)H:quinone oxidoreductase." PLoS One 7(8);e43902. PMID: 22952804
Lacour & Landini, 2004: Lacour S, Landini P (2004). "SigmaS-dependent gene expression at the onset of stationary phase in Escherichia coli: function of sigmaS-dependent genes and identification of their promoter sequences." J Bacteriol 186(21);7186-95. PMID: 15489429
Lasserre06: Lasserre JP, Beyne E, Pyndiah S, Lapaillerie D, Claverol S, Bonneu M (2006). "A complexomic study of Escherichia coli using two-dimensional blue native/SDS polyacrylamide gel electrophoresis." Electrophoresis 27(16);3306-21. PMID: 16858726
Liu04a: Liu X, De Wulf P (2004). "Probing the ArcA-P modulon of Escherichia coli by whole genome transcriptional analysis and sequence recognition profiling." J Biol Chem 279(13);12588-97. PMID: 14711822
LopezCampistrou05: Lopez-Campistrous A, Semchuk P, Burke L, Palmer-Stone T, Brokx SJ, Broderick G, Bottorff D, Bolch S, Weiner JH, Ellison MJ (2005). "Localization, annotation, and comparison of the Escherichia coli K-12 proteome under two states of growth." Mol Cell Proteomics 4(8);1205-9. PMID: 15911532
Natalello07: Natalello A, Doglia SM, Carey J, Grandori R (2007). "Role of flavin mononucleotide in the thermostability and oligomerization of Escherichia coli stress-defense protein WrbA." Biochemistry 46(2);543-53. PMID: 17209564
Noll06: Noll G, Kozma E, Grandori R, Carey J, Schodl T, Hauska G, Daub J (2006). "Spectroelectrochemical investigation of a flavoprotein with a flavin-modified gold electrode." Langmuir 22(5);2378-83. PMID: 16489832
Weichart03: Weichart D, Querfurth N, Dreger M, Hengge-Aronis R (2003). "Global role for ClpP-containing proteases in stationary-phase adaptation of Escherichia coli." J Bacteriol 185(1);115-25. PMID: 12486047
Wolfova07: Wolfova J, Mesters JR, Brynda J, Grandori R, Natalello A, Carey J, Kuta Smatanova I (2007). "Crystallization and preliminary diffraction analysis of Escherichia coli WrbA in complex with its cofactor flavin mononucleotide." Acta Crystallogr Sect F Struct Biol Cryst Commun 63(Pt 7);571-5. PMID: 17620713
Wolfova09: Wolfova J, Smatanova IK, Brynda J, Mesters JR, Lapkouski M, Kuty M, Natalello A, Chatterjee N, Chern SY, Ebbel E, Ricci A, Grandori R, Ettrich R, Carey J (2009). "Structural organization of WrbA in apo- and holoprotein crystals." Biochim Biophys Acta 1794(9);1288-98. PMID: 19665595
Yang93a: Yang W, Ni L, Somerville RL (1993). "A stationary-phase protein of Escherichia coli that affects the mode of association between the trp repressor protein and operator-bearing DNA." Proc Natl Acad Sci U S A 90(12);5796-800. PMID: 8516330
Zafar09: Zafar MN, Tasca F, Gorton L, Patridge EV, Ferry JG, Noll G (2009). "Tryptophan repressor-binding proteins from Escherichia coli and Archaeoglobus fulgidus as new catalysts for 1,4-dihydronicotinamide adenine dinucleotide-dependent amperometric biosensors and biofuel cells." Anal Chem 81(10);4082-8. PMID: 19438267
Zhang09a: Zhang J, Sprung R, Pei J, Tan X, Kim S, Zhu H, Liu CF, Grishin NV, Zhao Y (2009). "Lysine acetylation is a highly abundant and evolutionarily conserved modification in Escherichia coli." Mol Cell Proteomics 8(2);215-25. PMID: 18723842
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