Photosensitized oxidative DNA damage: From hole injection to chemical product formation and strand cleavage

Oxidatively generated damage to DNA induced by a pyrenyl photosensitizer residue (Py) covalently attached to a guanine base in the DNA sequence context 5′-d(CAT[G₁^Py]CG₂TCCTAC) in aerated solutions was monitored from the initial one-electron transfer, or hole injection step, to the formation of chemical end-products monitored by HPLC, mass spectrometry, and high-resolution gel electrophoresis. Hole injection into the DNA was initiated by two-photon excitation of the Py residue with 355 nm laser pulses, thus producing the radical cation Py^•+ and hydrated electrons; the latter are trapped by O₂, thus forming the superoxide anion O₂^•+. The decay of the Py ^•+ radical is correlated with the appearance of the G ^•+/G(-H)^• radical on microsecond time scales, and O₂^•- combines with guanine radicals at G ₁ to form alkali-labile 2,5-diamino-4H-imidazolone lesions (Iz ₁^Py). Product formation in the modified strand is smaller by a factor of 2.4 in double-stranded than in single-stranded DNA. In double-stranded DNA, hot piperidine-mediated cleavage at G₂ occurs only after G₁^Py, an efficient hole trap, is oxidized thus generating tandem lesions. An upper limit of hole hopping rates, k_hh < 5 × 10³ s^-1 from G₁ ^•+-Py to G₂ can be estimated from the known rates of the combination reaction of the G(-H)^• and O₂ ^•- radicals. The formation of Iz products in the unmodified complementary strand compared to the modified strand in the duplex is ∼10 times smaller. The formation of tandem lesions is observed even at low levels of irradiation corresponding to "single-hit" conditions when less than ∼10% of the oligonucleotide strands are damaged. A plausible mechanism for this observation is discussed.