Deamination of cytosine to uracil is one of the major events of DNA damage. Repair of damaged uracil DNA is achieved in a base excision repair pathway initiated by a Uracil DNA glycosylase (UDG). In present study, a DFT calculation was used to evaluate the potential energy surface and locate the correct transition state (TS) conformation between the connected reactant and product. Gaussian calculations including single-point energies calculations, geometry optimizations for reactant and transition state, frequency calculations, electrostatic potential calculations, and intrinsic reaction coordinate (IRC) calculations at the B3LYP/6-31+G(d) level of theory were performed. The transition state structures were successfully confirmed by the presence of only one imaginary frequency in vibration analysis and the corresponding mode motion is the cleavage stretching of the C-N glycosidic bond associated with bond cleavage. Our study showed that the change in hybridization from sp3 to sp2 on the anomeric carbon partially neutralizes the positive charge, and creates a partial empty 2pz orbital which hyperconjugates with the lone pair of O4’ and (C2’-H2’) electrons, thus stabilizing the transition state. A typical 3’-exo sugar ring conformation that favors the oxacarbenium ion through nucleotide backbone distortion was also identified. Our transition state model agrees well with the experimental results. These TS analyses have provided a detailed view of the mechanism of the hydrolytic deamination repair enzyme UDG.

Transition-state analysis of the uracil DNA glycosylase

251st National Meeting (2016)

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