Femtosecond Dynamics of Flavin Cofactor in DNA Photolyase:  Radical Reduction, Local Solvation, and Charge Recombination

We report here our femtosecond studies of the photoreduction dynamics of the neutral radical flavin (FADH) cofactor in E. coli photolyase, a process converting the inactive form to the biologically active one, a fully reduced deprotonated flavin FADH-. The observed temporal absorption evolution revealed two initial electron-transfer reactions, occurring in 11 and 42 ps with the neighboring aromatic residues of W382 and F366, respectively. The new transient absorption, observed at 550 nm previously in photolyase, was found from the excited-state neutral radical and is probably caused by strong interactions with the adenine moiety through the flavin U-shaped configuration and the highly polar/charged surrounding residues. The solvation dynamics from the locally ordered water molecules in the active site was observed to occur in ∼2 ps. These ultrafast ordered-water motions are critical to stabilizing the photoreduction product FADH- instantaneously to prevent fast charge recombination. The back electron-transfer reaction was found to occur in ∼3 ns. This slow process, consistent with ultrafast stabilization of the catalytic cofactor, favors photoreduction in photolyase.