MetaCyc Enzyme: cytosine deaminase

Gene: codA Accession Numbers: EG11326 (MetaCyc), b0337, ECK0334

Species: Escherichia coli K-12 substr. MG1655

Subunit composition of cytosine deaminase = [CodA]6

Cytosine deaminase (CDA) is an enzyme in the pyrimidine salvage pathway, catalyzing the deamination of cytosine to uracil, which allows the cell to utilize cytosine for pyrimidine nucleotide synthesis [deHaan72, Ahmad72]. It was recently shown to catalyze deamination of isoguanine, a mutagenic oxidation product of adenine [Hitchcock11, Friedman51].

As a mononuclear iron enzyme, CDA is damaged by micromolar hydrogen peroxide both in vitro and in vivo. Iron sequestration and substitution of Mn2+ for the Fe2+ cofactor can protect the enzyme [Anjem12].

Earlier characterization of the enzyme had found it to consist of two polypeptides of different molecular mass [Katsuragi86]. Later studies did not confirm this observation, finding only a single polypeptide [Danielsen92].

Crystal structures of wild-type and mutant CodA have been solved [Ireton01, Ireton02, Mahan04, Fuchita09, Hall11, Hitchcock11]. The enzyme is in a hexameric domain-swapped β-barrel conformation with the active site located at the barrel opening [Ireton02]. Random and site-directed mutagenesis has been used to identify active site residues and mutants that are more active with 5-fluorocytosine as substrate [Mahan04a, Mahan04, Fuchita09, Hall11]. A reaction mechanism has been proposed [Hall11].

Expression of CodA is repressed by the presence of pyrimidine nucleobases in the medium and increased by growth on poor nitrogen sources [Andersen89]. The transcription start site and subsequent elongation to a full codBA mRNA is regulated by the level of UTP [Qi95]. Regulation is reviewed in [Turnbough08].

Cytosine deaminase is present in prokaryotes and fungi, but not in multicellular eukaryotes. It is thus of interest for the design of antimicrobial agents. Cytosine deaminase is also being used for suicide gene therapy against tumors, e.g. [Mullen92, Yazawa02].

Citations: [Austin93, Reaves13]

Locations: cytosol

Map Position: [355,395 -> 356,678]

Molecular Weight of Polypeptide: 47.591 kD (from nucleotide sequence), 48.0 kD (experimental) [Danielsen92 ]

pI: 6.09

Unification Links: ASAP:ABE-0001161 , CGSC:913 , DIP:DIP-9306N , EchoBASE:EB1302 , EcoGene:EG11326 , EcoliWiki:b0337 , Mint:MINT-1307853 , ModBase:P25524 , OU-Microarray:b0337 , PortEco:codA , PR:PRO_000022316 , Pride:P25524 , Protein Model Portal:P25524 , RefSeq:NP_414871 , RegulonDB:EG11326 , SMR:P25524 , String:511145.b0337 , UniProt:P25524

Relationship Links: InterPro:IN-FAMILY:IPR011059 , InterPro:IN-FAMILY:IPR013108 , PDB:Structure:1K6W , PDB:Structure:1K70 , PDB:Structure:1R9X , PDB:Structure:1R9Y , PDB:Structure:1R9Z , PDB:Structure:1RA0 , PDB:Structure:1RA5 , PDB:Structure:1RAK , PDB:Structure:3G77 , PDB:Structure:3O7U , PDB:Structure:3R0D , PDB:Structure:3RN6 , Pfam:IN-FAMILY:PF07969

Gene-Reaction Schematic: ?

Gene-Reaction Schematic

GO Terms:

Biological Process: GO:0006209 - cytosine catabolic process Inferred from experiment [deHaan72]
GO:0019858 - cytosine metabolic process Inferred by computational analysis [UniProtGOA11a]
Molecular Function: GO:0004131 - cytosine deaminase activity Inferred from experiment Inferred by computational analysis [GOA01, Porter93]
GO:0008198 - ferrous iron binding Inferred from experiment [Porter93]
GO:0008270 - zinc ion binding Inferred from experiment [Hall11]
GO:0035888 - isoguanine deaminase activity Inferred from experiment [Hitchcock11]
GO:0042802 - identical protein binding Inferred from experiment [Rajagopala14, Lasserre06, Ireton02]
GO:0016787 - hydrolase activity Inferred by computational analysis [UniProtGOA11a]
GO:0016810 - hydrolase activity, acting on carbon-nitrogen (but not peptide) bonds Inferred by computational analysis [GOA01a]
GO:0046872 - metal ion binding Inferred by computational analysis [UniProtGOA11a]
Cellular Component: GO:0005829 - cytosol Inferred from experiment Inferred by computational analysis [DiazMejia09, Lasserre06]

MultiFun Terms: metabolism central intermediary metabolism nucleotide and nucleoside conversions

Created in EcoCyc 11-May-2011 by Keseler I , SRI International
Imported from EcoCyc 02-Jun-2015 by Paley S , SRI International

Enzymatic reaction of: cytosine deaminase

Synonyms: cytosine aminohydrolase

EC Number:

cytosine + H2O + H+ <=> uracil + ammonium

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 physiologically favored in the direction shown.

In Pathways: pyrimidine nucleobases salvage II , pyrimidine ribonucleosides salvage III

Imported from EcoCyc 02-Jun-2015 by Paley S , SRI International

Mn2+, Co2+, and Zn2+ can subsitute for Fe2+, but result in lower kcat values [Porter93].

Cofactors or Prosthetic Groups [Comment 1]: Fe2+ [Porter93]

Alternative Cofactors for Fe2+: Mn2+ , Co2+ , Zn2+

Inhibitors (Competitive): Cu2+ (Kic = 1.8µM) [Porter93, Comment 2] , Zn2+ (Kic = 5.8µM) [Porter93, Comment 3]

Inhibitors (Unknown Mechanism): hydrogen peroxide [Anjem12] , phosphonocytosine [Hall11] , o-phenanthroline [Porter93]

Kinetic Parameters:

Km (μM)
kcat (sec-1)
kcat/Km (sec-1 μM-1)

T(opt): 50 °C [Katsuragi86]

pH(opt): 9 [Katsuragi86]

Enzymatic reaction of: isoguanine deaminase (cytosine deaminase)

isoguanine + H+ + H2O <=> xanthine + ammonium

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 physiologically favored in the direction shown.

Imported from EcoCyc 02-Jun-2015 by Paley S , SRI International

The enzyme was purified from E. coli BL21(DE3), a B (not K-12) strain [Hitchcock11]. The protein sequences are identical.

Kinetic Parameters:

Km (μM)
kcat (sec-1)
kcat/Km (sec-1 μM-1)

Sequence Features

Feature Class Location Citations Comment
Cleavage-of-Initial-Methionine 1
UniProt: Removed;
Chain 2 -> 427
UniProt: Cytosine deaminase;
Extrinsic-Sequence-Variant 13
UniProt: In strain: SO5076..
Metal-Binding-Site 62
UniProt: Iron.
Metal-Binding-Site 64
UniProt: Iron.

10/20/97 Gene b0337 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG11326; confirmed by SwissProt match.


Ahmad72: Ahmad SI, Pritchard RH (1972). "Location of gene specifying cytosine deaminase in Escherichia coli." Mol Gen Genet 118(4);323-5. PMID: 4570159

Andersen89: Andersen L, Kilstrup M, Neuhard J (1989). "Pyrimidine, purine and nitrogen control of cytosine deaminase synthesis in Escherichia coli K 12. Involvement of the glnLG and purR genes in the regulation of codA expression." Arch Microbiol 152(2);115-8. PMID: 2673119

Anjem12: Anjem A, Imlay JA (2012). "Mononuclear iron enzymes are primary targets of hydrogen peroxide stress." J Biol Chem 287(19);15544-56. PMID: 22411989

Austin93: Austin EA, Huber BE (1993). "A first step in the development of gene therapy for colorectal carcinoma: cloning, sequencing, and expression of Escherichia coli cytosine deaminase." Mol Pharmacol 43(3);380-7. PMID: 8450832

CHARGAFF48: CHARGAFF E, KREAM J (1948). "Procedure for the study of certain enzymes in minute amounts and its application to the investigation of cytosine deaminase." J Biol Chem 175(2);993. PMID: 18880798

Danielsen92: Danielsen S, Kilstrup M, Barilla K, Jochimsen B, Neuhard J (1992). "Characterization of the Escherichia coli codBA operon encoding cytosine permease and cytosine deaminase." Mol Microbiol 1992;6(10);1335-44. PMID: 1640834

deHaan72: de Haan PG, Felix HS, Peters R (1972). "Mapping of the gene for cytosine deaminase on the Escherichia coli chromosome." Antonie Van Leeuwenhoek 38(3);257-63. PMID: 4561896

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

Friedman51: Friedman S, Gots JS (1951). "Deamination of isoguanine by Escherichia coli." Arch Biochem Biophys 32(1);227-9. PMID: 14847690

Fuchita09: Fuchita M, Ardiani A, Zhao L, Serve K, Stoddard BL, Black ME (2009). "Bacterial cytosine deaminase mutants created by molecular engineering show improved 5-fluorocytosine-mediated cell killing in vitro and in vivo." Cancer Res 69(11);4791-9. PMID: 19487291

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."

Hall11: Hall RS, Fedorov AA, Xu C, Fedorov EV, Almo SC, Raushel FM (2011). "Three-dimensional structure and catalytic mechanism of Cytosine deaminase." Biochemistry 50(22);5077-85. PMID: 21545144

Hitchcock11: Hitchcock DS, Fedorov AA, Fedorov EV, Dangott LJ, Almo SC, Raushel FM (2011). "Rescue of the orphan enzyme isoguanine deaminase." Biochemistry 50(25);5555-7. PMID: 21604715

Ireton01: Ireton GC, Black ME, Stoddard BL (2001). "Crystallization and preliminary X-ray analysis of bacterial cytosine deaminase." Acta Crystallogr D Biol Crystallogr 57(Pt 11);1643-5. PMID: 11679731

Ireton02: Ireton GC, McDermott G, Black ME, Stoddard BL (2002). "The structure of Escherichia coli cytosine deaminase." J Mol Biol 315(4);687-97. PMID: 11812140

Katsuragi86: Katsuragi T, Sakai T, Matsumoto K, Tonomura K (1986). "Cytosine deaminase from Escherichia coli - production, purification, and some characteristics." Agric Biol Chem 50(7):1721-1730.

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

Mahan04: Mahan SD, Ireton GC, Knoeber C, Stoddard BL, Black ME (2004). "Random mutagenesis and selection of Escherichia coli cytosine deaminase for cancer gene therapy." Protein Eng Des Sel 17(8);625-33. PMID: 15381761

Mahan04a: Mahan SD, Ireton GC, Stoddard BL, Black ME (2004). "Alanine-scanning mutagenesis reveals a cytosine deaminase mutant with altered substrate preference." Biochemistry 43(28);8957-64. PMID: 15248753

Mullen92: Mullen CA, Kilstrup M, Blaese RM (1992). "Transfer of the bacterial gene for cytosine deaminase to mammalian cells confers lethal sensitivity to 5-fluorocytosine: a negative selection system." Proc Natl Acad Sci U S A 89(1);33-7. PMID: 1729703

Porter00: Porter DJ (2000). "Escherichia coli cytosine deaminase: the kinetics and thermodynamics for binding of cytosine to the apoenzyme and the Zn(2+) holoenzyme are similar." Biochim Biophys Acta 2000;1476(2);239-52. PMID: 10669789

Porter93: Porter DJ, Austin EA (1993). "Cytosine deaminase. The roles of divalent metal ions in catalysis." J Biol Chem 1993;268(32);24005-11. PMID: 8226944

Qi95: Qi F, Turnbough CL (1995). "Regulation of codBA operon expression in Escherichia coli by UTP-dependent reiterative transcription and UTP-sensitive transcriptional start site switching." J Mol Biol 254(4);552-65. PMID: 7500333

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

Reaves13: Reaves ML, Young BD, Hosios AM, Xu YF, Rabinowitz JD (2013). "Pyrimidine homeostasis is accomplished by directed overflow metabolism." Nature 500(7461);237-41. PMID: 23903661

Turnbough08: Turnbough CL, Switzer RL (2008). "Regulation of pyrimidine biosynthetic gene expression in bacteria: repression without repressors." Microbiol Mol Biol Rev 72(2);266-300, table of contents. PMID: 18535147

UniProt09: UniProt Consortium (2009). "UniProt version 15.8 released on 2009-10-01 00:00:00." Database.

UniProt10: UniProt Consortium (2010). "UniProt version 2010-11 released on 2010-11-02 00:00:00." Database.

UniProt15: UniProt Consortium (2015). "UniProt version 2015-01 released on 2015-01-16 00:00:00." Database.

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

Yazawa02: Yazawa K, Fisher WE, Brunicardi FC (2002). "Current progress in suicide gene therapy for cancer." World J Surg 26(7);783-9. PMID: 11948367

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Please cite the following article in publications resulting from the use of MetaCyc: Caspi et al, Nucleic Acids Research 42:D459-D471 2014
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