Model studies of the (6-4) photoproduct photolyase enzyme: Laser flash photolysis experiments confirm radical ion intermediates in the sensitized repair of thymine oxetane adducts

The DNA (6-4) photoproduct photolyases are proteins that bind to UV-damaged DNA, specifically to sites that contain a (6-4) pyrimidine-pyrimidone lesion. Upon absorption of UV-A and visible light they catalyze the reversal of these lesions back to normal bases. It has been proposed that the photorepair occurs via an oxetane intermediate, which is formed from a ring-closing isomerization of the (6-4) photoproduct. Four model compounds for the oxetane intermediate have been prepared through photocycloaddition of carbonyl compounds (benzophenone, benzaldehyde, tolualdehyde, and anisaldehyde) with the 5,6 C=C of 1,3-dimethylthymine. The behavior of these compounds under sensitized photolysis conditions has been examined. On the basis of laser flash photolysis, fluorescence quenching, and product analysis experiments, it is demonstrated that these oxetane intermediates undergo a cycloreversion reaction upon photosensitized reductive electron-transfer reactions. The cycloreversion process yields the anion radicals of the carbonyl compounds. A lower limit on the rate constant of this anion radical splitting reaction is estimated to be >5 x 10(7) s(-1) These results support the proposed mechanism for DNA (6-4) photoproduct photolyase.