BINDING AND KINETICS STUDIES OF OXIDATION OF DNA BY OXORUTHENIUM(IV)

The binding to DNA of complexes based on Ru(tpy)(L)OH22+ (tpy = 2,2',2''-terpyridine; L = bpy, 2,2'-bipyridine; phen, 1,10-phenanthroline; or dppz, dipyridophenazine) has been studied by viscometry,thermal denaturation, and absorbance hypochromism along with the kinetics of oxidation of DNA by the analogous Ru(tpy)(L)O2+ complexes. These studies show that very weak binding occurs when L = bpy; however, when L = dppz, DELTAT(m) is larger than that for ethidium bromide. Viscometry studies of the dppz complex show that the dppz complex does lengthen DNA, as occurs with intercalative binding. The slope of the viscometry plot is identical to that for ethidium bromide, and neighbor exclusion binding is observed for both, with saturation occurring between 0.2 and 0.25 small molecules per nucleotide phosphate. The Ru(tpy)(dppz)OH22+ complex also unwinds DNA by 17 +/- 2., as determined using a topoisomerase assay. For L = bpy, no evidence of DNA lengthening was obtained. The kinetics of the oxidation of DNA by the Ru(IV)O2+ forms of these complexes occurs in two phases. The first phase involves oxidation of DNA by Ru(IV)O2+ that is bound at time zero. The second phase occurs when a reduced Ru(II)OH22+ complex dissociates and another Ru(IV)O2+ complex binds from solution. The kinetics of this phase are governed by dissociation of the reduced complex, which allows the relative dissociation rates of the L = bpy, phen, and dppz complexes to be determined. These experiments show that the dissociation rate for the dppz complex is an order of magnitude slower than those for bpy and phen, which is also consistent with an intercalative interaction for dppz. The cleavage reaction is shown to lead to the release of nucleic acid bases, implicating sugar oxidation as the reaction pathway.