| Abstract: | Endonuclease V of bacteriophage T4 has been described as an enzyme, coded for by the denV gene, that incises UV-irradiated DNA. It has recently been proposed that incision of irradiated DNA by this enzyme and the analogous "correndonucleases" I and II of Micrococcus luteus requires the sequential action of a pyrimidine dimer-specific DNA glycosylase and an apyrimidinic/apurinic endonuclease. In support of this two-step mechanism, we found that our preparations of T4 endonuclease V contained a DNA glycosylase activity that produced alkali-labile sites in irradiated DNA and an apyrimidinic/apurinic endonuclease activity that converted these sites to nicks. Both activities could be detected in the presence of 10 mM EDTA. In experiments designed to determine which of the activities is coded by the denV gene, we found that the glycosylase was more heat labile in extracts of Escherichia coli infected with either of two thermosensitive denV mutants than in extracts of cells infected with wild-type T4. In contrast, apyrimidinic/apurinic endonuclease activity was no more heat labile in extracts of the former than in extracts of the latter. Our results indicate that the denV gene codes for a DNA glycosylase specific for pyrimidine dimers. |
