Kinetic analysis of nucleotides incorporated opposite oxidized thymine bases on template DNA

Incorporation of nucleotides opposite the oxidized thymine bases on the template by eukaryotic DNA polymerases alpha and gamma was studied using the synthetic oligonucleotides containing 5-formyluracil or 5-hydroxymethyluracil. Primer extension assay revealed that both polymerases could bypass the oxidized thymine lesion, and incorporated dATP and dGTP opposite 5-formyluracil on the template. On the other hand, dATP was an only nucleotide that was incorporated opposite 5-hydroxymethyluracil by both polymerases as far as examined. These results indicate that 5-formyluracil but not 5-hydroxymethyluracil on template DNA may induce the transition mutation of A:T to G:C in eukaryotes.