Structural and biochemical characterization of yeast alkylpurine DNA glycosylases
Work presented in this dissertation details structural and biochemical characterization of three yeast alkylpurine DNA glycosylases – Mag1 and Mag2 from S. pombe and Mag from S. cerevisiae. I determined high resolution crystal structures all three enzymes and used the structures to initiate a deeper understanding of the sources of substrate specificity and catalytic competence in DNA glycosylases. My study of Mag1 and Mag provided novel evidence that protein-DNA interactions away from the active site can modulate the specificity of DNA glycosylases and a single substitution in the minor-groove interrogating loop is sufficient to alter the preference of one alkylpurine glycosylase for εA to that of a homologous enzyme. The crystal structure of Mag2 along with biochemical and phylogenetic analysis of the Mag genes in related fungal species studies showed that spMag2 cannot form a catalytically competent complex with DNA due to differences in the minor-groove interrogating loop and overall electrostatic surface potential compared to other alkylpurine glycosylases and may have evolved to perform a separate function.