| DNA lesions can be generated at an estimated rate of up to one million molecular lesions per cell per day, and are induced by a wide range of endogenous and exogenous damaging agents. These can include reactive oxygen species, alkylating agents, and genotoxic chemicals, all of which can yield lesion formation upon reaction with all four nucleobases of DNA. The mutagenic potential of a lesion rests on its intrinsic chemical structure, as well as the polymerase involved in its bypass. Many lesions distort DNA double helix, and are known blocks to replication. Stalled or blocked replication forks can trigger cell death and lead to genomic instability, so the cell has evolved numerous pathways to either repair or tolerate the damage in the event that it is unable to be excised and repaired. Given the wide range of damaging agents and resultant damaged bases, understanding their effects on replication can be complex, and requires the individual analysis of each lesion to determine its mutagenic potential.;During lesion bypass, the polymerase encounters two kinetic barriers, which coincides with the insertion opposite the lesion, as well as the subsequent extension. This dissertation monitors the kinetics of nucleotide insertion opposite and past four different types of lesions mediated by polymerases known to be important in lesion bypass, which include the Pols eta, iota, kappa, zeta, and gamma. The carboxymethylated lesions, cyclopurine lesions, ethylated thymidine lesions, and ribonucleotides embedded in DNA compromised the accuracy and efficiency of DNA replication to varying degrees. Most strikingly was the mutation profile generated from these replication studies closely matched the known mutation signature for several human diseases, including gastrointestinal cancers in the case of the carboxymethylated lesions. As a result, the studied lesions are attractive candidates for further research on their potential roles in the etiology of human diseases, and use as biomarkers for various ailments. |
