On the molecular basis of transition mutations. Frequency of forming 2-aminopurine-cytosine base mispairs in the G X C----A X T mutational pathway by T4 DNA polymerase in vitro

J Biol Chem. 1984 Oct 10;259(19):11713-7.


An in vitro model system including wild-type T4 DNA polymerase, the mutagenic nucleotide analogue 2-aminopurine deoxyribonucleoside triphosphate, and poly[d(A,C)] X oligo(dT) poly(dC) X oligo(dG) template-primers is used to measure the frequency of 2-aminopurine X cytosine base mispairs formed in the G X C----A X T mutational pathway. Incorporation and turnover of the analogue into DNA is dependent on the presence of cytosine on the template strand and is reduced significantly in the presence of dGTP. 2-Aminopurine X cytosine mispairs are observed to occur at a 2-3 order of magnitude greater frequency than adenine X cytosine mispairs. The frequency of inserting 2-aminopurine deoxyribonucleoside monophosphate in place of dGMP opposite template cytosine sites is about 3-6% when either strong or weak base-stacking partners are present on the primer strand. However, enzymatic proofreading of the mispair strongly depends on base-stacking partners. Greater than 85% of misinserted 2-aminopurine deoxynucleotides are excised whenever the mispairs are formed next to 5'-primer thymine sites. A 5-fold reduction in proofreading frequency occurs when the mispair is formed with 2-aminopurine deoxynucleoside monophosphate stacked adjacent to a 5'-primer guanine. The frequency of 2-aminopurine X cytosine base mispair formation in the G X C----A X T pathway is similar to that found previously in the A X T----G X C pathway (Watanabe, S. M., and Goodman, M.F. (1981) Proc. Natl. Acad. Sci. U.S.A. 78, 2864-2868). We propose a criterion for base selection by DNA polymerase to account for the unexpected similarity in base mispairing rates in the two transition pathways.

Publication types

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

MeSH terms

  • 2-Aminopurine / analysis*
  • Adenine / analogs & derivatives*
  • Base Sequence
  • Cytosine / analysis*
  • DNA-Directed DNA Polymerase / metabolism*
  • Mutation*
  • Polydeoxyribonucleotides / metabolism
  • T-Phages / enzymology*
  • Templates, Genetic
  • Time Factors


  • Polydeoxyribonucleotides
  • poly(dA-dC).oligo(dT)
  • 2-Aminopurine
  • Cytosine
  • DNA-Directed DNA Polymerase
  • Adenine