DB code: D00675

RLCP classification 5.201.2781500.563 : Elimination
4.12.642320.562 : Addition
CATH domain 2.30.40.10 : Urease, subunit C; domain 1
3.20.20.140 : TIM Barrel Catalytic domain
E.C. 3.5.4.1
CSA
M-CSA
MACiE

CATH domain Related DB codes (homologues)
3.20.20.140 : TIM Barrel S00231 S00232 M00186 D00673 D00801 D00873 M00030 M00225 M00226
2.30.40.10 : Urease, subunit C; domain 1 D00673 D00801 D00873 M00030 M00225 M00226

Uniprot Enzyme Name
UniprotKB Protein name Synonyms RefSeq Pfam
P25524 Cytosine deaminase
EC 3.5.4.1
Cytosine aminohydrolase
NP_414871.1 (Protein)
NC_000913.2 (DNA/RNA sequence)
YP_488631.1 (Protein)
NC_007779.1 (DNA/RNA sequence)
PF07969 (Amidohydro_3)
[Graphical View]

KEGG enzyme name
Cytosine deaminase
Isocytosine deaminase

UniprotKB: Accession Number Entry name Activity Subunit Subcellular location Cofactor
P25524 CODA_ECOLI Cytosine + H(2)O = uracil + NH(3). Homotetramer. Iron or another divalent metal ion.

KEGG Pathways
Map code Pathways E.C.
MAP00240 Pyrimidine metabolism

Compound table
Cofactors Substrates Products Intermediates
KEGG-id C14818 C00380 C00001 C00106 C00014 I00151
E.C.
Compound Fe2+ cytosine H2O uracil NH3 4-Hydroxy-cytosine
Type heavy metal amide group,amine group,aromatic ring (with nitrogen atoms) H2O amide group,aromatic ring (with nitrogen atoms) amine group,organic ion
ChEBI 29033
16040
15377
17568
16134
PubChem 27284
597
22247451
962
1174
222
1k6wA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound Unbound
1k70A01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound Unbound
1r9xA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound Unbound
1r9yA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound Unbound
1r9zA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound Unbound
1ra0A01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound Unbound
1ra5A01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound Unbound
1rakA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound Unbound
3g77A01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound Unbound
3o7uA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound Unbound
3r0dA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound Unbound
3rn6A01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound Unbound
1k6wA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_FE Unbound Unbound Unbound Unbound
1k70A02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_FE Unbound Unbound Unbound Intermediate-analogue:HPY
1r9xA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_FE Unbound Unbound Unbound Unbound
1r9yA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_FE Unbound Unbound Unbound Unbound
1r9zA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_FE Unbound Unbound Unbound Unbound
1ra0A02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_FE Unbound Unbound Unbound Intermediate-analogue:FPY
1ra5A02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_FE Unbound Unbound Unbound Intermediate-analogue:FPY
1rakA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_FE Unbound Unbound Unbound Intermediate-analogue:FPY
3g77A02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_FE Unbound Unbound Unbound Unbound
3o7uA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Analogue:_ZN Unbound Unbound Unbound Intermediate-analogue:O7U
3r0dA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Analogue:_ZN Unbound Unbound Unbound Unbound
3rn6A02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Analogue:_ZN Analogue:IGA Unbound Unbound Unbound

Reference for Active-site residues
resource references E.C.
PDB;1k6w, 1k70 & literature [6]

Active-site residues
PDB Catalytic residues Cofactor-binding residues Modified residues Main-chain involved in catalysis Comment
1k6wA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain
1k70A01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain
1r9xA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain
1r9yA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain
1r9zA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain
1ra0A01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain
1ra5A01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain
1rakA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain
3g77A01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain
3o7uA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain
3r0dA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain
3rn6A01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain
1k6wA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain GLU 217;HIS 246;ASP 313 HIS 61;HIS 63;HIS 214;ASP 313(Fe2+ binding)
1k70A02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain GLU 217;HIS 246;ASP 313 HIS 61;HIS 63;HIS 214;ASP 313(Fe2+ binding)
1r9xA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain GLU 217;HIS 246;ASP 313 HIS 61;HIS 63;HIS 214;ASP 313(Fe2+ binding) mutant D314G
1r9yA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain GLU 217;HIS 246;ASP 313 HIS 61;HIS 63;HIS 214;ASP 313(Fe2+ binding) mutant D314A
1r9zA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain GLU 217;HIS 246;ASP 313 HIS 61;HIS 63;HIS 214;ASP 313(Fe2+ binding) mutant D314S
1ra0A02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain GLU 217;HIS 246;ASP 313 HIS 61;HIS 63;HIS 214;ASP 313(Fe2+ binding) mutant D314G
1ra5A02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain GLU 217;HIS 246;ASP 313 HIS 61;HIS 63;HIS 214;ASP 313(Fe2+ binding) mutant D314A
1rakA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain GLU 217;HIS 246;ASP 313 HIS 61;HIS 63;HIS 214;ASP 313(Fe2+ binding) mutant D314S
3g77A02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain GLU 217;HIS 246;ASP 313 HIS 61;HIS 63;HIS 214;ASP 313(Fe2+ binding) mutant V152A, F316C, D317G
3o7uA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain GLU 217;HIS 246;ASP 313 HIS 61;HIS 63;HIS 214;ASP 313(Fe2+ binding)
3r0dA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain GLU 217;HIS 246;ASP 313 HIS 61;HIS 63;HIS 214;ASP 313(Fe2+ binding)
3rn6A02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain GLU 217;HIS 246;ASP 313 HIS 61;HIS 63;HIS 214;ASP 313(Fe2+ binding)

References for Catalytic Mechanism
References Sections No. of steps in catalysis
[6]
p. 693-694
[12]
Scheme 4

References
[1]
Resource
Comments
Medline ID
PubMed ID 1925539
Journal Science
Year 1991
Volume 252
Pages 1278-84
Authors Wilson DK, Rudolph FB, Quiocho FA
Title Atomic structure of adenosine deaminase complexed with a transition-state analog: understanding catalysis and immunodeficiency mutations.
Related PDB
Related UniProtKB
[2]
Resource
Comments
Medline ID
PubMed ID 8439534
Journal Biochemistry
Year 1993
Volume 32
Pages 1689-94
Authors Wilson DK, Quiocho FA
Title A pre-transition-state mimic of an enzyme: X-ray structure of adenosine deaminase with bound 1-deazaadenosine and zinc-activated water.
Related PDB
Related UniProtKB
[3]
Resource
Comments
Medline ID
PubMed ID 8226944
Journal J Biol Chem
Year 1993
Volume 268
Pages 24005-11
Authors Porter DJ, Austin EA
Title Cytosine deaminase. The roles of divalent metal ions in catalysis.
Related PDB
Related UniProtKB
[4]
Resource
Comments
Medline ID
PubMed ID 9622483
Journal Biochemistry
Year 1998
Volume 37
Pages 8314-24
Authors Wang Z, Quiocho FA
Title Complexes of adenosine deaminase with two potent inhibitors: X-ray structures in four independent molecules at pH of maximum activity.
Related PDB
Related UniProtKB
[5]
Resource
Comments
Medline ID
PubMed ID 10669789
Journal Biochim Biophys Acta
Year 2000
Volume 1476
Pages 239-52
Authors Porter DJ
Title Escherichia coli cytosine deaminase: the kinetics and thermodynamics for binding of cytosine to the apoenzyme and the Zn(2+) holoenzyme are similar.
Related PDB
Related UniProtKB
[6]
Resource
Comments X-RAY CRYSTALLOGRAPHY (1.75 ANGSTROMS).
Medline ID
PubMed ID 11812140
Journal J Mol Biol
Year 2002
Volume 315
Pages 687-97
Authors Ireton GC, McDermott G, Black ME, Stoddard BL
Title The structure of Escherichia coli cytosine deaminase.
Related PDB 1k6w 1k70
Related UniProtKB P25524
[7]
Resource
Comments X-RAY CRYSTALLOGRAPHY (1.6 ANGSTROMS).
Medline ID
PubMed ID 12637534
Journal J Biol Chem
Year 2003
Volume 278
Pages 19111-7
Authors Ko TP, Lin JJ, Hu CY, Hsu YH, Wang AH, Liaw SH
Title Crystal structure of yeast cytosine deaminase. Insights into enzyme mechanism and evolution.
Related PDB
Related UniProtKB
[8]
Resource
Comments X-RAY CRYSTALLOGRAPHY (1.14 ANGSTROMS) IN COMPLEX WITH SUBSTRATE ANALOG AND ZINC IONS, AND SUBUNIT.
Medline ID
PubMed ID 12906827
Journal Structure
Year 2003
Volume 11
Pages 961-72
Authors Ireton GC, Black ME, Stoddard BL
Title The 1.14 A crystal structure of yeast cytosine deaminase: evolution of nucleotide salvage enzymes and implications for genetic chemotherapy.
Related PDB
Related UniProtKB
[9]
Resource
Comments
Medline ID
PubMed ID 15468078
Journal Angew Chem Int Ed Engl
Year 2004
Volume 43
Pages 5396-9
Authors Sponer JE, Sanz Miguel PJ, Rodriguez-Santiago L, Erxleben A, Krumm M, Sodupe M, Sponer J, Lippert B
Title Metal-mediated deamination of cytosine: experiment and DFT calculations.
Related PDB
Related UniProtKB
[10]
Resource
Comments
Medline ID
PubMed ID 15248753
Journal Biochemistry
Year 2004
Volume 43
Pages 8957-64
Authors Mahan SD, Ireton GC, Stoddard BL, Black ME
Title Alanine-scanning mutagenesis reveals a cytosine deaminase mutant with altered substrate preference.
Related PDB
Related UniProtKB
[11]
Resource
Comments X-RAY CRYSTALLOGRAPHY.
Medline ID
PubMed ID 15381761
Journal Protein Eng Des Sel
Year 2004
Volume 17
Pages 625-33
Authors Mahan SD, Ireton GC, Knoeber C, Stoddard BL, Black ME
Title Random mutagenesis and selection of Escherichia coli cytosine deaminase for cancer gene therapy.
Related PDB 1r9x 1r9y 1r9z 1ra0 1ra5 1rak
Related UniProtKB
[12]
Resource
Comments
Medline ID
PubMed ID 21545144
Journal Biochemistry
Year 2011
Volume 50
Pages 5077-85
Authors Hall RS, Fedorov AA, Xu C, Fedorov EV, Almo SC, Raushel FM
Title Three-dimensional structure and catalytic mechanism of cytosine deaminase.
Related PDB 3o7u
Related UniProtKB
[13]
Resource
Comments
Medline ID
PubMed ID 21604715
Journal Biochemistry
Year 2011
Volume 50
Pages 5555-7
Authors Hitchcock DS, Fedorov AA, Fedorov EV, Dangott LJ, Almo SC, Raushel FM
Title Rescue of the orphan enzyme isoguanine deaminase.
Related PDB 3rn6
Related UniProtKB

Comments
This enzyme is homologous to adenosine deaminase (EC 3.5.4.4; S00232 in EzCatDB) and guanine deaminase (EC 3.5.4.3; D00873 in EzCatDB).
Although its catalytic site is similar to those of the homologous enzymes, cofactor ion, Fe2+, is different from the cofactor ion, zinc ion, adopted by the homologous enzymes. However, the reaction mechanism seems to be very similar to those homologous enzymes.
According to the literature [12] and the reaction mechanism of the homologous enzymes(S00232 and D00873 in EzCatDB), this enzyme catalyzes two successive reactions (rather than hydrolysis) as follows:
(A) Addition of water to imine carbon to form a tetrahedral intermediate (I00151).
(A1) Asp313 (or Glu217) acts as a general base to deprotonate the Fe2+-bound water. Here, the positive charge of His246 seems to stabilize the activated water. His246 may assist the catalytic function of Glu217 as well (see [12]).
(A2) The activated water makes a nucleophilic attack on the C4 atom of cytosine, whilst Glu217 acts as a general acid to protonate the N3 atom (protonation site) of the cytosine. This reaction leads to the formation of tetrahedral intermediate at the C4 atom, transforming N3-C4 bond from a double bond to a single bond (I00151).
(B) Elimination of amine group from the intermediate (I00151), forming a carbonyl group.
(B1) Asp313 acts as a general acid to protonate the eliminated amine group, releasing the ammonia.
(B2) Asp313 acts as a general base to deprotonate the hydroxyl group, bound to the Fe2+ ion and His246. (Here, His246 may assist the catalytic function of Asp313 as well.) This reaction leads to the enol form of the product, cytosine. (This may be an E1-like reaction, as amine elimination occurs prior to deprotonation.)

Created Updated
2008-05-30 2012-10-17