DNA damage by chemically generated singlet oxygen.
A naphthalenic endoperoxide was used as a non-photochemical source of singlet oxygen (1O2) to examine some interactions between this reactive oxygen species and DNA. High molecular weight DNA (ca. 10(8) daltons) was exposed to 120 mol m-3 1O2 (cumulative concentration) and analyzed for interstrand crosslinkage by hydroxyl apatite chromatography following formamide denaturation. No evidence for 1O2-induced interstrand crosslinking was obtained. The capacity of 1O2 to generate strand breaks in single-stranded (ss) and double-stranded (ds) DNA was investigated by sucrose gradient centrifugation analysis of bacteriophage phi X174 DNA. No direct strand breaks could be detected at neutral pH, whereas extensive strand breakage was observed after treatment with alkali. Possible biological consequences of 1O2-exposure were assessed by examining the plaque-forming capacity of ss and ds phi X174 DNA molecules using wildtype Escherichia coli spheroplasts as recipients. Without any further treatment with heat or alkali, exposure to the endoperoxide resulted in a time- and dose-dependent inactivation, ss DNA being considerably more sensitive than ds DNA. From the present results and those reported earlier (Nieuwint et al.,) we infer that 1O2-induced inactivation of phi X174 DNA is not due to DNA backbone breakage nor to interstrand crosslinking, but rather to some form of damage to the base or sugar moiety of the DNA, the exact nature of which remains to be elucidated.
Lafleur
M V
MV
Department of Biophysics, Free University, Amsterdam, The Netherlands.
Nieuwint
A W
AW
Aubry
J M
JM
Kortbeek
H
H
Arwert
F
F
Joenje
H
H
D010584:Bacteriophage phi X 174 / Q000235:genetics; D002499:Centrifugation, Density Gradient; D003432:Cross-Linking Reagents; D004247:DNA / Q000187:drug effects; D004249:DNA Damage; D004279:DNA, Viral / Q000187:drug effects; D003903:Deuterium; D017666:Deuterium Oxide; D009284:Naphthols / Q000494:pharmacology; D010100:Oxygen / Q000494:pharmacology*; D014867:Water
D003432:Cross-Linking Reagents; D004279:DNA, Viral; D009284:Naphthols; D014867:Water; D004247:DNA; C058008:3,3'-(1,4-naphthylidene)diproprionate; D003903:Deuterium; D017666:Deuterium Oxide; D010100:Oxygen