Redox reactivity of guanyl radicals in plasmid DNA

Abstract

Purpose : It has been previously argued that γ-irradiation of plasmid DNA in the presence of thiocyanate ions produces products recognized by the E. coli base excision-repair endonuclease formamidopyrimidine-DNA N -glycosylase (FPG), and there that derive from an intermediate guanyl radical species. The wish was to characterize the reactivity of this intermediate with reducing agents. Materials and methods : Aqueous solutions of plasmid DNA containing either bromide or thiocyanate (10 -3 to 10 -1 mol dm -3) and also one of six other additives (azide, ferrocyanide, iodide, nitrite, promethazine, tryptophan, 10 -7 to 10 -3 mol dm -3) were subjected to 137 Cs γ-irradiation (662 keV). After irradiation, the plasmid was incubated with FPG. Strand break yields before and after incubation were determined by agarose gel electrophoresis under neutral conditions. Results : The very high yields of FPG-sensitive sites in the presence of SCN - or Br - decreased significantly with increasing concentrations of all of the six additives, with promethazine and tryptophan being the most efficient additives, and azide and iodide the least. Conclusions : From the results it is possible to estimate values of the rate constants for the reduction of the DNA guanyl radical (5x10 5, 2x10 5, 10 7 and 10 7 dm 3 mol -1 s -1 for ferrocyanide, nitrite, promethazine and tryptophan respectively).