Yield of DNA strand breaks after base oxidation of plasmid DNA

We have irradiated aerobic aqueous solutions of plasmid DNA with Cs-137 gam ma rays in the presence of inorganic radical scavengers including nitrite, iodide, azide, thiocyanate and bromide. These scavengers react with the str ongly oxidizing hydroxyl radical ((OH)-O-.) to produce less powerful oxidan ts. Of these scavengers, only thiocyanate and bromide result in the formati on of oxidizing species [(SCN)(2)(.-) and Br-2(.-), respectively] which are capable of reacting with the bases in DNA. The oxidized bases were detecte d after incubation of the irradiated plasmid with the two E. coli DNA base excision repair endonucleases, formamidopyrimidine-DNA N-glycosylase and en donuclease III. Depending on the experimental conditions, the intermediate base radicals mag ultimately form stable oxidized bases in very high yields (within an order of magnitude of the (OH)-O-. yield), and possibly also si ngle-strand breaks (SSBs) in much lower yield (between 0.1 and 1% of the to tal yield of base damage). By competing for (SCN)(2)(.-) with an additional species (nitrite), it was possible to estimate the second-order rate const ant for the reaction of (SCN)(2)(.-) with DNA as 1.6 x 10(4) dm(3) mol(-1) s(-1), and also to demonstrate a correlation between the large Field of dam aged bases and the much smaller increase in the yield of SSBs over backgrou nd levels due to (OH)-O-.. The efficiency of transfer of damage from oxidiz ed base to sugar is estimated as about 0.5% or 5%, depending on whether pur ine or pyrimidine base radicals are responsible for the base to sugar damag e transfer. (C) 1999 by Radiation Research Society.