Ultraviolet solar radiation may induce DNA damage that can lead to skin cancer.[1] Cyclobutane pyrimidine dimers (CPDs) and pyrimidine-pyrimidone (6-4) photoproducts ((6-4)PPs) are two of the most abundant cytotoxic, mutagenic and carcinogenic DNA photolesions.[2] To maintain genetic integrity, many organisms have developed an efficient mechanism to repair these lesions by near-UV/blue light driven enzymes called DNA photolyases.[3] Compared to the nearly fully identified CPD photorepair, the repair mechanism of (6-4)PPs remains elusive, including lesion recognition and accommodation, photoinduced electron transfer to the lesion, the splitting of the pyrimidine dimer, and restoration of the original bases. Base on classical molecular dynamics (MD) simulations, we investigate the lesion recognition process by characterising the energetic properties and key intermediate states. The protonation states of key species in the reaction centre, such as His365 and His 369, are highly controversial whereas crucial for the proton transfer process during repair. To address this, combined quantum mechanics/molecular mechanics (QM/MM) simulations along with pKa calculations are carried out to assign the protonation states. The role of nearby protein residues and water molecules in the binding pocket is also probed by analysing the energy profiles of different reaction pathways obtained from QM/MM simulations. The contributions from key residues are studied through perturbative analysis based on the wild-type calculations by switching off their electrostatic interactions.