Hydrolysis of the damaged deoxythymidine glycol nucleoside and comparison to canonical DNA.

Genomic integrity is continually under attack by both endogenous and exogenous sources. One of the most common forms of damage is oxidation of the thymine nucleobase to form (5R,6S)-dihydroxy-5,6-dihydro-thymine (thymine glycol or Tg), which stops DNA polymerases and is thus cytotoxic. Thymine glycol damage is repaired through a variety of mechanisms, including the multi-step base excision repair (BER) pathway. In the first BER step, the glycosidic bond of the dTg nucleotide is hydrolyzed by a DNA glycosylase. In order to understand the catalytic effect of the glycosylases, the corresponding uncatalyzed mechanisms and barriers are required, as well as an appreciation of the relative reactivity of the glycosidic bond with respect to the corresponding canonical nucleoside. To this end, the PCM-B3LYP/6-31+G(d) reaction potential energy surfaces (PES) for deoxythymidine (dT) and dTg hydrolysis are characterized in the present study using solvent-phase optimizations and a model containing three explicit water molecules. The surfaces are comparable to those generated using functionals that account for dispersion interactions (B3LYP-D3 and M06-2X). Mapping the PES as a function of the glycosidic bond length and nucleophile-sugar distance reveals a synchronous S(N)2 mechanism as the lowest energy pathway for damaged dTg hydrolysis, which contrasts the preferred dissociative S(N)1 mechanism isolated for the deglycosylation of natural dT. As proposed for other enzymes, the difference in excision pathway may at least in part help the enzyme selectively target the damaged base and discriminate against the natural counterpart. Interestingly, the barrier to dTg deglycosylation (ΔG(‡) = 138.0 kJ mol(-1)) is much higher than for dT deglycosylation (ΔG(‡) = 112.7 kJ mol(-1)), which supports the stability of this lesion and clarifies the catalytic feat presented to DNA repair enzymes that remove this detrimental damage from the genome. Although nucleotide excision repair (NER) typically targets bulky DNA lesions, the large calculated barrier for dTg deglycosylation rationalizes why the NER mechanism also excises this non-bulky lesion from cellular DNA.