Proteins of the Int family and proteins of the topoisomerase I family have a fundamentally similar biochemical mechanism; they catalyze reversible breakage and joining of DNA by forming a covalent intermediate in which a tyrosine residue of the protein is covalently joined to DNA. Although the covalent intermediate is designed to be transient, accidents occur in which it is stabilized; the resulting long-lived break in DNA can threaten the survival of the affected cell. We recently discovered an enzyme, a tyrosine-DNA phosphodiesterase, that is ubiquitous in eukaryotes (but apparently absent in bacteria) and can unlink a complete topoisomerase as well as an isolated tyrosine moiety from dead-end complexes with DNA. We have now partially purified the activity from the yeast, S. cerevisiae, and have characterized its biochemical properties. We have shown that the enzyme has a Km for tyrosine-DNA adducts in the nanomolar range, indicating that the protein readily accepts its putative natural substrate. By studying classic phosphodiesterase substrates, we have proven that the enzyme is hydrolytic and have confirmed its strong preference for tyrosine linked to the 3' end of DNA as opposed to that linked to the 5' end of DNA. Taken together with its broad distribution in the animal kingdom, the specificity and robustness of the enzyme lead us to conclude that it is likely to play a significant role in the DNA repair of topoisomerase-induced breaks in the genome.
