Effect of β,γ-CHF- and β,γ-CHCl-dGTP halogen atom stereochemistry on the transition state of DNA polymerase β

Biochemistry. 2012 Oct 30;51(43):8491-501. doi: 10.1021/bi3010335. Epub 2012 Oct 19.

Abstract

Recently, we synthesized the first individual β,γ-CHX-dGTP diastereomers [(R)- or (S)-CHX, where X is F or Cl] and determined their structures in ternary complexes with DNA polymerase β (pol β). We now report stereospecificity by pol β on the mixed β,γ-CHX diastereomer pairs using nuclear magnetic resonance and on the separate diastereomers using transient kinetics. For both the F and Cl diastereomers, the R isomer is favored over the S isomer for G·C correct incorporation, with stereospecificities [(k(pol)/K(d))(R)/(k(pol)/K(d))(S)] of 3.8 and 6.3, respectively, and also for G·T misincorporation, with stereospecificities of 11 and 7.8, respectively. Stereopreference for the (R)-CHF-dGTP diastereomer was abolished for k(pol) but not K(d) with mutant pol β (R183A). These compounds constitute a new class of stereochemical probes for active site interactions involving halogen atoms. As Arg183 is unique in family X pols, the design of CXY deoxyribonucleotide analogues to enhance interaction is a possible strategy for inhibiting BER selectively in cancer cells.

Publication types

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

MeSH terms

  • Catalytic Domain / drug effects
  • DNA / metabolism
  • DNA Polymerase beta / chemistry
  • DNA Polymerase beta / genetics
  • DNA Polymerase beta / metabolism*
  • Deoxyguanine Nucleotides / chemistry*
  • Deoxyguanine Nucleotides / pharmacology*
  • Halogens / chemistry*
  • Halogens / pharmacology*
  • Humans
  • Kinetics
  • Nuclear Magnetic Resonance, Biomolecular
  • Point Mutation
  • Stereoisomerism
  • Substrate Specificity

Substances

  • Deoxyguanine Nucleotides
  • Halogens
  • deoxyguanosine triphosphate
  • DNA
  • DNA Polymerase beta