N7-alkyl-guanine (N7aG) has long been well known as a major type of DNA lesion that could lead to various cancers. However, due to the difficulty of site-specific incorporation, their cytotoxic and mutagenic effects, and particularly the in vivo repairing mechanisms have not been well studied. It has been suggested that these lesions can be repaired through base excision repair (BER) pathway which involves the recognition and removal of the damaged bases by glycosylase, hydrolysis by endonuclease and recruitment of one of the most important BER repairing proteins: DNA polymerase � (Pol�).  In addition to the aforementioned significances, N7aG lesions are interesting in themselves because of two more reasons: 1) different bulky adduct groups could affect the nucleotide conformations resulting in duplex structural or base pairing changes; 2) N7 atom is positively charged in a way that has been suggested changing the charge distribution of the base ring decreasing the pKa of the N1-hydrogen atom, which could lead to mismatch. To further explore the effects of N7aG lesions, we applied with the methods of crystallography and kinetics. We first further extended the glycosidic bond stabilizing method of adding 2'-flourination on the deoxyribose moiety, and were able to synthesize scalable amounts of N7-methyl, benzyl, naphtalenyl Guanine phosphoramidites for site-specific incorporation using ultra-mild DNA synthesis method. With these in hand, we explored the insertions and pairing of these lesions in the DNA sequences by Pol�, the protein crucial in the BER.

Structural and kinetic explorations of N7-alkyl Guanine lesions

247th National Meeting (2014)

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