Translesional synthesis on DNA templates containing 8-oxo-7,8-dihydrodeoxyadenosine

This study was designed to establish the miscoding potential of 8-oxo-7,8-dihydrodeoxyadenosine (8-oxo-dA). Oligodeoxynucleotides modified site-specifically with 8-oxo-dA were used as templates in primer extension reactions catalyzed by DNA polymerase I (Klenow fragment), DNA polymerase alpha (pol alpha), or DNA polymerase beta (pol beta). dTMP or dGMP is incorporated opposite 8-oxo-dA when either of these dNTPs is provided as substrate for DNA polymerase. dTMP is incorporated exclusively opposite 8-oxo-dA when all four dNTPs are present in the reaction mixture at equimolar concentrations. Chain extension is catalyzed efficiently by Klenow fragment and pol beta under conditions where 8-oxo-dA is paired with dT at the 3' terminus of the primed DNA template. Chain extension catalyzed by pol alpha proceeds more slowly. As shown by steady-state kinetic experiments, incorporation of dGMP is higher in reactions catalyzed by pol beta than by Klenow fragment or pol alpha. The dG-8-oxo-dA pair is extended efficiently from the 3' terminus in the absence of dTTP. We conclude that DNA containing 8-oxo-dA is capable of miscoding; however, unlike 8-oxo-dG, the mutagenic potential of this lesion is limited.