UV-induced pyrimidine hydrates in DNA are repaired by bacterial and mammalian DNA glycosylase activities.
Escherichia coli endonuclease III and mammalian repair enzymes cleave UV-irradiated DNA at AP sites formed by the removal of cytosine photoproducts by the DNA glycosylase activity of these enzymes. Poly(dG-[3H]dC) was UV irradiated and incubated with purified endonuclease III. 3H-Containing material was released in a fashion consistent with Michaelis-Menten kinetics. This 3H material was determined to be cytosine by chromatography in two independent systems and microderivatization. 3H-Containing material was not released from nonirradiated copolymer. When poly(dA-[3H]dU) was UV irradiated, endonuclease III released 3H-containing material that coeluted with uracil hydrate (6-hydroxy-5,6-dihydrouracil). Similar results are obtained by using extracts of HeLa cells. There results indicate that the modified cytosine residue recognized by endonuclease III and the mammalian enzyme is cytosine hydrate (6-hydroxy-5,6-dihydrocytosine). Once released from DNA through DNA-glycosylase action, the compound eliminates water, reverting to cytosine. This is consistent with the known instability of cytosine hydrate. The repairability of cytosine hydrate in DNA suggests that it is stable in DNA and potentially genotoxic.
Boorstein
R J
RJ
Department of Pathology, New York University Medical Center, New York 10016.
Hilbert
T P
TP
Cadet
J
J
Cunningham
R P
RP
Teebor
G W
GW
D000818:Animals; D003596:Cytosine / Q000032:analysis; D045647:DNA Glycosylases; D004260:DNA Repair; D043224:Deoxyribonuclease (Pyrimidine Dimer); D004706:Endodeoxyribonucleases / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology; D029968:Escherichia coli Proteins; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D013058:Mass Spectrometry; D009699:N-Glycosyl Hydrolases / Q000378:metabolism*; D011089:Polydeoxyribonucleotides / Q000138:chemical synthesis / Q000528:radiation effects*; D014316:Tritium; D014466:Ultraviolet Rays
D029968:Escherichia coli Proteins; D011089:Polydeoxyribonucleotides; D014316:Tritium; D003596:Cytosine; D004706:Endodeoxyribonucleases; D043224:Deoxyribonuclease (Pyrimidine Dimer); C106403:NTH protein, E coli; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases