Reaction of guanyl radicals in plasmid DNA with biological reductants: chemical repair of DNA damage produced by the direct effect of ionizing radiation

Purpose: It has been previously argued that the use of the one-electron oxi dants (SCN)(2)(.-) and Br-2(.-) with plasmid DNA leads to the formation of DNA guanyl radicals. These guanyl radical species are intermediates in the DNA damage produced by processes such as photo-ionization and ionizing irra diation. The present paper evaluates the use of thallium(II) ions ((TlOH+)- O-II) as the one-electron oxidant, and also determines rate constants for t he reduction (repair) of guanyl radicals in plasmid DNA by a variety of red ucing agents including the biologically important compounds ascorbate and g lutathione. Materials and methods: Aqueous solutions of plasmid DNA containing 10(-3) m ol dm(-3) thiocyanate or thallous ions and a reducing agent (azide, nitrite , ferrocyanide, hexachloroiridate( III), iodide, ascorbate, glutathione, gl utathione disulphide, methionine, tyrosine, 5-hydroxyindole-3-acetic acid, 10(-7)-10(-4) mol dm(-3)) were irradiated with Cs-137 gamma -rays (662 keV) . After irradiation, the plasmid was incubated with the E. coli base excisi on repair endonuclease formamidopyrimidine-DNA N-glycosylase (FPG). Strand break yields after incubation were quantified by means of agarose gel elect rophoresis. Results: High yields of FPG-sensitive sites produced by the oxi dants (SCN)(2)(.-) and (TlOH+)-O-II were strongly attenuated by the presenc e of the reducing agents. Conclusions : From the results, it is possible to arrive at estimates of th e rate constants for the reduction of the DNA guanyl radical by the reducin g agents. Values lie in the range 10(4)-10(7) dm(3) mol(-1) s(-1). Using th e values for ascorbate and glutathione, it is possible to estimate an upper limit on the order of milliseconds for the lifetime of DNA guanyl radicals under cellular conditions. The implication is that there may well be a sig nificant chemical repair of DNA base damage by the direct effect of ionizin g radiation.