Hypoxanthine Excision from DNA in Escherichia Coli and Saccharomyces Cerevisiae
Defective temporal regulation of some DNA repair enzymes, including hypoxanthine-DNA glycosylase is in part responsible for Bloom's syndrome. Identification of this enzyme in yeast would provide a simple eukaryotic system in which to study its function and regulation. An attempt was made to identify hypoxanthine-DNA glycosylase activity in Saccharomyces cerevisiae. Wild type (wt) and uracil-DNA glycosylase deficient (ung-) strains of both E. coli and S. cerevisiae were transformed with double-stranded phagemids pRS316 and pRS314 prepared from single-stranded normal (obtained from a wild type strain) and uracil-containing DNA (U-DNA). The second strand of each was synthesized to contain various ratios of inosine:guanine and the second strand of U-DNA was also synthesized with different ratios of uracil:thymine and transformed into both strains of each species as a control. Wild typeE. coli exhibited a pattern of more transform ants at lower I:G ratios even when background numbers of transformants, from incomplete degradation of the uracil strand, were taken into account. This suggests that hypoxanthine-DNA glycosylase is inefficient at cleaving I -C base pairs. The sustained ability of inosine-uracil hybrids to transform both wt and ung- S. cerevisiae strains at wild type levels, after degradation of the uracil strand suggests that hypoxanthine-DNA glycosylase in yeast is inefficient at cleaving I-C base pairs, if it is present at all.