Figure 3.

Functional analyses of residues in the AAG active site. Wild-type and mutant AAG proteins were expressed in a S. cerevisiae strain lacking the endogeneous Mag1 glycosylase activity, and resistance to the alkylating agent methylmethane sulfonate was determined by growth on a gradient of MMS. The extent of growth on MMS is shown for cells expressing a basal level of AAG (gray) and induced cells expressing a higher level of AAG protein (black). Immunoblots indicated that all of the AAG mutant proteins are expressed at the same level as the wild-type AAG protein (not shown). Compared with the null cells (−AAG), cells expressing wild-type AAG (wt) are resistant to growth inhibition by MMS. Substitution of Glu-125 with alanine (E125A) or glutamine (E125Q) eliminates detectable MMS resistance. The functional significance of these residues is discussed in the text.