Polymerization of 2'-fluoro- and 2'-O-methyl-dNTPs by human DNA polymerase alpha, polymerase gamma, and primase

Biochem Pharmacol. 2000 May 1;59(9):1045-52. doi: 10.1016/s0006-2952(99)00414-1.


Studies were undertaken to assess the ability of human polymerase alpha (pol alpha) and polymerase gamma (pol gamma) to incorporate 2'-fluoro- and 2'-O-methyldeoxynucleotides into DNA. In vitro DNA synthesis systems were used to detect incorporation and determine K(m) and V(max) for 2'-FdATP, 2'-FdUTP, 2'-FdCTP, 2'-FdGTP, 2'-O-MedATP, 2'-O-MedCTP, 2'-O-MedGTP, 2'-O-MedUTP, dUTP, UTP, and FIAUTP, in addition to normal deoxynucleotides. Pol alpha incorporated all 2'-FdNTPs except 2'-FdATP, but not 2'-O-MedNTPs. Pol gamma incorporated all 2'-FdNTPs, but not 2'-O-MedNTPs. In general, 2'-fluorine substitution decreased V(max)/K(m) 2'-FdUTP. Because kinetics of insertion of pol alpha can be affected by the nature of the primer, we examined the ability of pol alpha to polymerize 2'-fluoro- and 2'-O-MedATP and dGTP when elongating a primer synthesized by DNA primase. Under these conditions, both 2'-FdATP and 2'-FdGTP were polymerized, but 2'-O-MedATP and 2'-O-MedGTP were not. Primase alone could not readily polymerize these analogs into RNA primers. Previous studies showed that 2'-deoxy-2'-fluorocytosine (2'-FdC) is incorporated by several non-human DNA polymerases. The current studies showed that human polymerases can polymerize numerous 2'-FdNTPs but cannot polymerize 2'-O-MedNTPs.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • DNA Polymerase I / metabolism*
  • DNA Polymerase gamma
  • DNA Primase / metabolism*
  • DNA-Directed DNA Polymerase / metabolism*
  • Deoxycytosine Nucleotides / chemistry
  • Deoxycytosine Nucleotides / metabolism
  • Deoxyguanine Nucleotides / chemistry
  • Deoxyguanine Nucleotides / metabolism
  • Deoxyribonucleotides / chemistry
  • Deoxyribonucleotides / metabolism*
  • Humans
  • Thymine Nucleotides / chemistry
  • Thymine Nucleotides / metabolism
  • Uridine Triphosphate / chemistry
  • Uridine Triphosphate / metabolism


  • Deoxycytosine Nucleotides
  • Deoxyguanine Nucleotides
  • Deoxyribonucleotides
  • Thymine Nucleotides
  • 2'-deoxycytidine 5'-triphosphate
  • deoxyguanosine triphosphate
  • DNA Primase
  • DNA Polymerase I
  • DNA Polymerase gamma
  • DNA-Directed DNA Polymerase
  • thymidine 5'-triphosphate
  • Uridine Triphosphate