A DNA oligonucleotide-hemin complex cleaves t-butyl hydroperoxide through a homolytic mechanism

Both electron paramagnetic resonance (EPR) and electronic absorption spectr oscopy have been employed to investigate the reaction of a guanine-rich DNA nucleotide-hemin complex (PS2.M-hemin complex) and organic peroxide (t-Bu- OOH). Incubation of the PS2.M-hemin complex with t-Bu-OOH resulted in the t ime-dependent decrease in the heme Soret with concomitant changes to the vi sible bands of the electronic absorbance spectrum for the PS2.M-hemin compl ex. Parallel EPR studies using the spin trap 5,5-dimethyl-1-pyrroline N-oxi de (DMPO) combined with spectral simulation demonstrated the presence of te rt-butyloxyl, carbon-centered methyl, and methyl peroxyl radicals as well a s a simple nitroxide (triplet) signal. Experiments, performed by maintainin g a constant ratio of t-Bu-OOH/PS2.M-hemin complex (similar to 35 mol/mol) while varying DMPO concentration, indicated that the relative contributions of each radical adduct to the composite EPR spectrum were significantly in fluenced by the DMPO concentration. For example, at DMPO/PS2.M-hemin of 10- 50 mol/mol, a complex mixture of radicals was consistently detected, wherea s at high trapping efficiency (i.e., DMPO/PS2.M-hemin of similar to 250 mol /mol) the tert-butyloxyl-DMPO adduct was predominant. In contrast, at relat ively low DMPO/PS2.M-hemin complex ratios of less than or equal to5 mol/mol , a simple nitroxide three-line EPR signal was detected largely in the abse nce of all other radicals. Together, these data indicate that tert-butyloxy l radical is the primary radical likely formed from the homolytic cleavage of the O-O peroxy bond of t-Bu-OOH, while methyl and methyl peroxyl radical s result from beta -scission of the primary tert-butyloxyl radical product.