Correlation between sequence-dependent glycosylase repair and the thermal stability of oligonucleotide duplexes containing 1,N6-ethenoadenine

Previous experiments on DNA sequence context reported that base modification, replication, and repair are affected by the nature of neighbor bases. We now report that repair by mammalian alkylpurine-DNA-N-glycosylases (APNG) of 15-mer oligonucleotides with a central 1,N-6-ethenoadenine (epsilon A), flanked by 5' and 3' tandem bases, is also highly sequence dependent. Oligonucleotides with the central sequences -GG epsilon AGG- or -CC epsilon ACC- are repaired 3-5-fold more efficiently than those containing -AA epsilon AAA- or -TT epsilon ATT- when using human or mouse APNG, Melting curves of the same duplexes showed that oligomers with G.C/C.G neighbors were less denatured than those with A.T/T.A neighbors at 37 degrees C. This sequence-dependent difference in denaturation correlates with the relative thermodynamic stability of oligomers with G.C/C.G or A.T/T.A neighbors. The dependence of repair on thermal stability was confirmed by enzyme reactions performed over 0-45 degrees C. Under these conditions, repair of epsilon A flanked by G.C/C.G was dramatically increased at 37 degrees C with continuous increase up to 45 degrees C, in contrast to that with flanking A.T/T.A pairs, which was in agreement with the degree of denaturation of these duplexes, These results indicate that the thermodynamic stability conferred by base pairs flanking epsilon A plays an essential role in maintaining the integrity of the duplex structure which is necessary for repair.