Characterization of the Escherichia coli x-ray endonuclease, endonuclease III

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Abstract
The X-ray endonuclease, endonuclease III of E. coli was purified to apparent homogeneity by using the criterion of sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The most purified fraction shows endonucleolytic activity against apurinic and apyrimidinic (AP) sites and a dose-dependent response to DNA that was X-irradiated, UV-irradiated or treated with OsO4. The endonuclease also nicks OsO4-treated DNA that was subsequently treated with alkali to produce fragmented thymine residues and DNA treated with potassium permanganate. The enzyme does not incise the alkali-labile sites present in DNA X-irradiated in vitro in the presence of hydroxyl radical scavengers. The most purified fractions exhibit 2 distinct activities, an AP endonuclease that cleaves on the 3'' side of the damage leaving a 3''-OH and a 5''-PO4 and a DNA N-glycosylase that recognizes at least 2 substrates, thymine glycol residues and urea residues. The glycosylase activity is sensitive to N-ethylmaleimide, while the AP endonuclease is not.