Hydrogen-bonding capability of a templating difluorotoluene nucleotide residue in an RB69 DNA polymerase ternary complex

J Am Chem Soc. 2011 Jul 6;133(26):10003-5. doi: 10.1021/ja2021735. Epub 2011 Jun 15.


Results obtained using 2,4-difluorotoluene nucleobase (dF) as a nonpolar thymine isostere by Kool and colleagues challenged the Watson-Crick dogma that hydrogen bonds between complementary bases are an absolute requirement for accurate DNA replication. Here, we report crystal structure of an RB69 DNA polymerase L561A/S565G/Y567A triple mutant ternary complex with a templating dF opposite dTTP at 1.8 Å-resolution. In this structure, direct hydrogen bonds were observed between: (i) dF and the incoming dTTP, (ii) dF and residue G568 of the polymerase, and (iii) dF and ordered water molecules surrounding the nascent base pair. Therefore, this structure provides evidence that a templating dF can form novel hydrogen bonds with the incoming dTTP and with the enzyme that differ from those formed with a templating dT.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Crystallography, X-Ray
  • DNA-Directed DNA Polymerase / chemistry*
  • DNA-Directed DNA Polymerase / genetics
  • DNA-Directed DNA Polymerase / metabolism
  • Hydrogen Bonding
  • Models, Molecular
  • Mutation
  • Protein Structure, Tertiary
  • Thymine Nucleotides / chemistry*
  • Toluene / analogs & derivatives*
  • Toluene / chemistry
  • Viral Proteins / chemistry*
  • Viral Proteins / genetics
  • Viral Proteins / metabolism


  • Thymine Nucleotides
  • Viral Proteins
  • Toluene
  • difluorotoluene
  • DNA-Directed DNA Polymerase
  • bacteriophage RB69 DNA polymerase
  • thymidine 5'-triphosphate