Thymine Dimer Photoreversal in Purine-Containing Trinucleotides

Cyclobutane pyrimidine dimer yields in UV-irradiated DNA are controlled by the equilibrium between forward and reverse photoreactions. Past studies have shown that dimer yields are suppressed at sites adjacent to a purine base, but the underlying causes are unclear. In order to investigate whether this suppression is the result of repair by electron transfer from a neighboring nucleobase, the yields and dynamics of the reverse reaction were studied using trinucleotides containing a cis-syn dimer (T <> T) flanked on the 5' or the 3' side by adenine or guanine. The probability of forming an excited state on T <> T or on the purine base was varied by tuning the irradiation wavelength between 240 and 280 nm. Cleavage quantum yields decrease by an order of magnitude over this wavelength range and are less than 1% at 280 nm, a wavelength that excites the purine base with more than 95% probability. Conditional quantum yields of cleavage for the trinucleotides given excitation of T <> T are similar in magnitude to the quantum yield of cleavage of unmodified T <> T. These results indicate that within experimental uncertainty all photoreversal in these single-stranded substrates is the result of direct electronic excitation of T <> T. Photolyase-like repair of T <> T due to electron transfer from an adjacent purine is negligible in these substrates. Instead, the observed variation in photoreversal quantum yields for adenine-versus guanine-flanked cis-syn dimer could be due to uncertainties in absorption cross sections or to a modest quenching effect by the purine on the excited state of T <> T. Pump probe measurements reveal that the excited-state lifetimes of A or G in the dimer-containing trinucleotides are unperturbed by the neighboring dimer, indicating that electron transfer from purine base to T <> T is not competitive with rapid excited-state deactivation. Pump probe measurements on unmodified T <> T in aqueous solution indicate that cleavage is most likely complete on a picosecond or subpicosecond time scale.