The structures of two intermediates en route to DNA repair within a (6-4) photolyase

Pyrimidine (6-4) pyrimidone photoproducts (6-4PPs) are a type of mutagenic DNA lesion caused by UV light crosslinking of adjacent pyrimidine bases. These lesions need to be repaired to preserve the integrity of the genome, and this task is handled by (6-4) photolyase enzymes. The repair process is intricate, involving the movement of a hydroxyl group from one base to another, possibly accompanied by or followed by the breaking of a bond between the bases.

However, the precise mechanism by which the enzyme carries out this reaction is still not fully understood. Traditionally, expert models propose a stepwise process where an oxetane intermediate is formed, but this has never been confirmed directly despite decades of study. By utilizing dose-controlled X-ray crystallography on (6-4) photolyase from Vibrio cholerae, the transformation of 6-4PP into an oxetane species was observed.

Additionally, an unanticipated intermediate featuring a hemi-cleaved state was identified. The stepwise progression from 6-4PP to oxetane, hemi-cleaved, and fully repaired species suggests a need to reconsider the current models explaining DNA repair by (6-4) photolyases.