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PDBsum entry 1ffl

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protein links
Transferase PDB id
1ffl
Jmol
Contents
Protein chain
264 a.a. *
Waters ×10
* Residue conservation analysis
PDB id:
1ffl
Name: Transferase
Title: Crystal structure of the apo-thymidylate synthase r166q muta
Structure: Thymidylate synthase. Chain: a. Engineered: yes. Mutation: yes
Source: Escherichia coli. Organism_taxid: 562. Expressed in: escherichia coli. Expression_system_taxid: 562. Gene.
Biol. unit: Dimer (from PDB file)
Resolution:
2.94Å     R-factor:   0.171     R-free:   0.220
Authors: R.R.Sotelo-Mundo,L.Changchien,F.Maley,W.R.Montfort
Key ref: R.R.Sotelo-Mundo et al. (2006). Crystal structures of thymidylate synthase mutant R166Q: structural basis for the nearly complete loss of catalytic activity. J Biochem Mol Toxicol, 20, 88-92. PubMed id: 16615077
Date:
25-Jul-00     Release date:   28-Jul-00    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chain
Pfam   ArchSchema ?
P0A884  (TYSY_ECOLI) -  Thymidylate synthase
Seq:
Struc:
264 a.a.
264 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 2 residue positions (black crosses)

 Enzyme reactions 
   Enzyme class: E.C.2.1.1.45  - Thymidylate synthase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]

      Pathway:
Folate Coenzymes
      Reaction: 5,10-methylenetetrahydrofolate + dUMP = dihydrofolate + dTMP
5,10-methylenetetrahydrofolate
+ dUMP
= dihydrofolate
+ dTMP
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     cytoplasm   1 term 
  Biological process     methylation   5 terms 
  Biochemical function     transferase activity     4 terms  

 

 
    reference    
 
 
J Biochem Mol Toxicol 20:88-92 (2006)
PubMed id: 16615077  
 
 
Crystal structures of thymidylate synthase mutant R166Q: structural basis for the nearly complete loss of catalytic activity.
R.R.Sotelo-Mundo, L.Changchien, F.Maley, W.R.Montfort.
 
  ABSTRACT  
 
Thymidylate synthase (TS) catalyzes the folate-dependent methylation of deoxyuridine monophosphate (dUMP) to form thymidine monophosphate (dTMP). We have investigated the role of invariant arginine 166, one of four arginines that contact the dUMP phosphate, using site-directed mutagenesis, X-ray crystallography, and TS from Escherichia coli. The R166Q mutant was crystallized in the presence of dUMP and a structure determined to 2.9 A resolution, but neither the ligand nor the sulfate from the crystallization buffer was found in the active site. A second structure determined with crystals prepared in the presence of dUMP and the antifolate 10-propargyl-5,8-dideazafolate revealed that the inhibitor was bound in an extended, nonproductive conformation, partially occupying the nucleotide-binding site. A sulfate ion, rather than dUMP, was found in the nucleotide phosphate-binding site. Previous studies have shown that the substitution at three of the four arginines of the dUMP phosphate-binding site is permissive; however; for Arg166, all the mutations lead to a near-inactive mutant. The present structures of TS R166Q reveal that the phosphate-binding site is largely intact, but with a substantially reduced affinity for phosphate, despite the presence of the three remaining arginines. The position of Cys146, which initiates catalysis, is shifted in the mutant and resides in a position that interferes with the binding of the dUMP pyrimidine moiety.