Theoretical Insights into N‑Glycoside Bond Cleavage of 5‑Carboxycytosine by Thymine DNA Glycosylase: A QM/MM Study

Thymine DNA glycosylase (TDG)-mediated excision of 5-formylcytosine
and 5-carboxylcytosine (5-caC) is a critical step in active DNA demethylation.
Herein, we employed a combined quantum mechanics/molecular mechanics
approach to investigate the reaction mechanism of TDG-catalyzed N-glycosidic
bond cleavage of 5-caC. The calculated results show that TDG-catalyzed
5-caC excision follows a concerted (SN2) mechanism in which
glycosidic bond dissociation is coupled with nucleophile attack. Protonation
of the 5-caC anion contributes to the cleavage of the N-glycoside
bond, in which the N3-protonated zwitterion and imino tautomers are
more favorable than carboxyl-protonated amino tautomers. This is consistent
with the experimental data. Furthermore, our results reveal that the
configuration rearrangement process of the protonated 5-caC would
lower the stability of the N-glycoside bond and substantially reduce
the barrier height for the subsequent C1′–N1 bond cleavage.
This should be attributed to the smaller electrostatic repulsion between
the leaving base and the negative phosphate group as a result of the
structural rearrangement.