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PDBsum entry 1ra0

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protein ligands metals links
Hydrolase PDB id
1ra0
Jmol PyMol
Contents
Protein chain
423 a.a. *
Ligands
FPY
Metals
_FE
Waters ×466
* Residue conservation analysis
PDB id:
1ra0
Name: Hydrolase
Title: Bacterial cytosine deaminase d314g mutant bound to 5-fluoro- hydroxy-3,4-dihydropyrimidine.
Structure: Cytosine deaminase. Chain: a. Synonym: cytosine aminohydrolase. Engineered: yes. Mutation: yes
Source: Escherichia coli. Organism_taxid: 562. Gene: coda, b0337. Expressed in: escherichia coli. Expression_system_taxid: 562.
Biol. unit: Hexamer (from PDB file)
Resolution:
1.12Å     R-factor:   0.168     R-free:   0.172
Authors: S.D.Mahan,G.C.Ireton,B.L.Stoddard,M.E.Black
Key ref: S.D.Mahan et al. (2004). Random mutagenesis and selection of Escherichia coli cytosine deaminase for cancer gene therapy. Protein Eng Des Sel, 17, 625-633. PubMed id: 15381761
Date:
31-Oct-03     Release date:   05-Oct-04    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chain
Pfam   ArchSchema ?
P25524  (CODA_ECOLI) -  Cytosine deaminase
Seq:
Struc:
427 a.a.
423 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 1 residue position (black cross)

 Enzyme reactions 
   Enzyme class: E.C.3.5.4.1  - Cytosine deaminase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: Cytosine + H2O = uracil + NH3
Cytosine
+ H(2)O
=
uracil
Bound ligand (Het Group name = FPY)
matches with 88.89% similarity
+ NH(3)
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     cytosol   1 term 
  Biological process     cytosine metabolic process   2 terms 
  Biochemical function     isoguanine deaminase activity     9 terms  

 

 
    Added reference    
 
 
Protein Eng Des Sel 17:625-633 (2004)
PubMed id: 15381761  
 
 
Random mutagenesis and selection of Escherichia coli cytosine deaminase for cancer gene therapy.
S.D.Mahan, G.C.Ireton, C.Knoeber, B.L.Stoddard, M.E.Black.
 
  ABSTRACT  
 
Cytosine deaminase (CD) is currently being used as a suicide gene for cancer gene therapy. The premise of this therapy is the preferential deamination of 5-fluorocytosine (5FC) to 5-fluorouracil by cancer cells expressing cytosine deaminase. However, a lack of efficient gene transfer to tumors combined with inefficient 5FC turnover currently limits the clinical applications of this gene therapy approach. We have used random mutagenesis to create novel bacterial cytosine deaminases that demonstrate an increased preference for 5FC over cytosine. Among the 15 mutants isolated, one conferred sensitivity to Escherichia coli in a negative selection system at a concentration of 5FC that was 10-fold lower than a sublethal dose for wild-type CD. Evaluation of individual substitutions found in this double mutant (Q102R, D314G) demonstrated that the substitution at residue D314 was solely responsible for the observed increase in sensitivity to 5FC. Additional mutagenesis at D314 resulted in the identification of two more substitutions with the ability to confer enhanced 5FC sensitivity to E.coli. Structure determinations of the three CD variants in the presence and absence of a transition state 5FC analogue provide insights to the determinants of substrate binding specificity at the 5' position of the pyrimidine ring. CD mutant D314A is a promising candidate for further gene therapy studies.
 

Literature references that cite this PDB file's key reference

  PubMed id Reference
21192992 B.F.Canine, Y.Wang, W.Ouyang, and A.Hatefi (2011).
Development of targeted recombinant polymers that can deliver siRNA to the cytoplasm and plasmid DNA to the cell nucleus.
  J Control Release, 151, 95.  
21381910 W.F.Lai (2011).
In vivo nucleic acid delivery with PEI and its derivatives: current status and perspectives.
  Expert Rev Med Devices, 8, 173-185.  
20817124 Y.Wang, B.F.Canine, and A.Hatefi (2011).
HSV-TK/GCV cancer suicide gene therapy by a designed recombinant multifunctional vector.
  Nanomedicine, 7, 193-200.  
20622857 G.J.Bartlett, A.Choudhary, R.T.Raines, and D.N.Woolfson (2010).
n-->pi* interactions in proteins.
  Nat Chem Biol, 6, 615-620.  
20179680 P.Li, Y.Liu, J.Maynard, Y.Tang, and A.Deisseroth (2010).
Use of adenoviral vectors to target chemotherapy to tumor vascular endothelial cells suppresses growth of breast cancer and melanoma.
  Mol Ther, 18, 921-928.  
19820714 P.H.Ear, and S.W.Michnick (2009).
A general life-death selection strategy for dissecting protein functions.
  Nat Methods, 6, 813-816.  
19100795 W.F.Lai, and M.C.Lin (2009).
Nucleic acid delivery with chitosan and its derivatives.
  J Control Release, 134, 158-168.  
17301844 C.P.Guise, J.I.Grove, E.I.Hyde, and P.F.Searle (2007).
Direct positive selection for improved nitroreductase variants using SOS triggering of bacteriophage lambda lytic cycle.
  Gene Ther, 14, 690-698.  
17495948 S.A.Kaliberov, J.M.Market, G.Y.Gillespie, V.Krendelchtchikova, D.Della Manna, J.C.Sellers, L.N.Kaliberova, M.E.Black, and D.J.Buchsbaum (2007).
Mutation of Escherichia coli cytosine deaminase significantly enhances molecular chemotherapy of human glioma.
  Gene Ther, 14, 1111-1119.  
17420581 Y.Hamaji, M.Fujimori, T.Sasaki, H.Matsuhashi, K.Matsui-Seki, Y.Shimatani-Shibata, Y.Kano, J.Amano, and S.Taniguchi (2007).
Strong enhancement of recombinant cytosine deaminase activity in Bifidobacterium longum for tumor-targeting enzyme/prodrug therapy.
  Biosci Biotechnol Biochem, 71, 874-883.  
17094807 J.H.Xu, and J.Messing (2006).
Maize haplotype with a helitron-amplified cytidine deaminase gene copy.
  BMC Genet, 7, 52.  
The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time.

 

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