Impact of thymine glycol damage on DNA duplex energetics: Correlations with lesion-induced biochemical and structural consequences

The magnitude and nature of lesion-induced energetic perturbations empirically correlate with mutagenicity/cytotoxicity profiles and can be predictive of lesion outcomes during polymerase-mediated replication in vitro. In this study, we assess the sequence and counterbase-dependent energetic impact of the Thymine glycol (T_g) lesion on a family of deoxyoligonucleotide duplexes. T_g damage arises from thymine and methyl-cytosine exposure to oxidizing agents or radiation-generated free-radicals. The T_g lesion blocks polymerase-mediated DNA replication in vitro and the unrepaired site elicits cytotoxic lethal consequences in vivo. Our combined calorimetric and spectroscopic characterization correlates T_g-induced energetic perturbations with biological and structural properties. Specifically, we incorporate a 5R-T_g isomer centered within the tridecanucleotide sequence 5′-GCGTACXCATGCG-3′ (X = T_g or T) which is hybridized with the corresponding complementary sequence 5′-CGCATGNGTACGC-3′ (N = A, G, T, C) to generate families of T_g-damaged (T_g·N) and lesion-free (T·N) duplexes. We demonstrate that the magnitude and nature of the T_g destabilizing impact is dependent on counterbase identity (i.e., A ∼ G<T<C). The observation that a T_g lesion is less destabilizing when positioned opposite purines suggests that favorable counterbase stacking interactions may partially compensate lesion-induced perturbations. Moreover, the destabilizing energies of T_g·N duplexes parallel their respective lesion-free T·N mismatch counterparts (i.e., G<T<C). Elucidation of T_g-induced destabilization relative to the corresponding undamaged mismatch energetics allows resolution of lesion-specific and sequence-dependent impacts. The T_g-induced energetic perturbations are consistent with its replication blocking properties and may serve as differential recognition elements for discrimination by the cellular repair machinery.