Novel inhibition of archaeal family-D DNA polymerase by uracil

Nucleic Acids Res. 2013 Apr;41(7):4207-18. doi: 10.1093/nar/gkt083. Epub 2013 Feb 13.

Abstract

Archaeal family-D DNA polymerase is inhibited by the presence of uracil in DNA template strands. When the enzyme encounters uracil, following three parameters change: DNA binding increases roughly 2-fold, the rate of polymerization slows by a factor of ≈ 5 and 3'-5' proof-reading exonuclease activity is stimulated by a factor of ≈ 2. Together these changes result in a significant decrease in polymerization activity and a reduction in net DNA synthesis. Pol D appears to interact with template strand uracil irrespective of its distance ahead of the replication fork. Polymerization does not stop at a defined location relative to uracil, rather a general decrease in DNA synthesis is observed. 'Trans' inhibition, the slowing of Pol D by uracil on a DNA strand not being replicated is also observed. It is proposed that Pol D is able to interact with uracil by looping out the single-stranded template, allowing simultaneous contact of both the base and the primer-template junction to give a polymerase-DNA complex with diminished extension ability.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • DNA / biosynthesis
  • DNA / chemistry
  • DNA / metabolism
  • DNA, Single-Stranded / chemistry
  • DNA, Single-Stranded / metabolism
  • DNA-Directed DNA Polymerase / metabolism*
  • Deoxyadenine Nucleotides / metabolism
  • Deoxyribonucleotides / metabolism
  • Exonucleases / metabolism
  • Nucleic Acid Synthesis Inhibitors
  • Pyrococcus / enzymology*
  • Templates, Genetic
  • Uracil / metabolism*

Substances

  • DNA, Single-Stranded
  • Deoxyadenine Nucleotides
  • Deoxyribonucleotides
  • Nucleic Acid Synthesis Inhibitors
  • Uracil
  • 2'-deoxy-5'-adenosine monophosphate
  • DNA
  • DNA-Directed DNA Polymerase
  • Exonucleases