The Role of Hydroxyl Radical Quenching in the Protection by Acetate and Ethylenediaminetetraacetate of Supercoiled Plasmid DNA from Ionizing Radiation-induced Strand Breakage

Abstract

A supercoiled plasmid of 7300 base pairs was isolated and exposed in various aqueous environments to 60Co gamma-radiation. Conversion of the supercoiled form to the relaxed circular and linear forms was monitored by agarose gel electrophoresis and quantified by fluorescence scanning of the gel. Acetate, which has been reported to affect the conformation of DNA in solution, decreased the radiosensitivity of the supercoil in a concentration-dependent manner. Acetate, formate, and azide anions, as well as mannitol, all protected the supercoil from relaxation in approximate proportion to the rate at which their solutions quench the hydroxyl radical. At concentrations greater than 300 mmol dm-3, however, the efficiency of acetate radioprotection is reduced. Disodium ethylenediaminetetraacetate protected the supercoil more efficiently than would be expected from the published value of its rate constant for quenching the hydroxyl radical.

Reaction rate coefficients of hydroxyl radical-induced DNA single- and α-type double-strand breaks

“…Some progress has been made in the research of the rate constant of • OH radicals inducing DNA SSBs. Generally, this rate constant, always depending on the respective research group, increases along with the scavenging capacity [10,11,12]. However, minor impurities (i.e., metal ions) in the DNA solution may affect the precision of measurements so that different rate constants might even be yielded in different research periods [10,13].…”

Formation of single- and double-strand breaks of pBR322 plasmid irradiated in the presence of scavengers

“…In the present investigation, we adapted the pBR322 DNA in vitro model system to study irradiation-induced SSB and DSB in the presence of TE (10 -2 mol dm -3 Tris and 10 -3 mol dm -3 ethylene diamine tetra-acetic acid, EDTA) and mannitol, whose reaction rate coefficient k s with • OH radical was reported to be 1.8×10 9 mol -1 dm 3 s -1 [20]. We analyzed the relationship between SSB and αDSB and that between α and β where α is the number of DSB per unit dose induced in one irradiation event and β is the number of DSB per unit squared dose induced by the combination of two independent SSB.…”

Impact of gamma irradiation on the transformation efficiency for extracellular plasmid DNA

“…Gamma irradiation generates hydroxyl radicals which cause DNA damage in aqueous solution. The chelating agent disodium ethylenediaminetetraacetate (EDTA), which was contained in the TE samples, is a quencher of hydroxyl radicals (Blazek and Peak, 1988;Tomita et al, 1995). The higher transformation efficiency of the TE samples compared with that of the freshwater samples could be due to the action of EDTA as a chemical radioprotector.…”