Light-activated signal transduction proteins use a series of isomerization and proton transfer reactions to trigger photo-motility and biological responses and generate a wide range of structural signalings.To provide a comprehensive mechanism of the overall process at the atomic level,we apply a CASPT2//CASSCF/AMBER QM/MM protocol to investigate the relaxation pathways for a variety of possible isomerization and proton transfer reactions upon the photoexcitation of the wild-type photoactive yellow protein(PYP)and S65T/H148D mutant of green fluorescent protein(GFP).The nonadiabatic relay of CI(S1/S0)conical intersection of PYP is unveiled to play the decisive role to bifurcate the excited state relaxation towards the complete and short photocycles.Two major and one minor deactivation channels were found starting from the CI(S1/S0)like intermediate of IT,producing the cis isomers pR1,ICP,and ICT through the hula twist,bicycle pedal and one-bond flip isomerization reactions.The overall photocycle of PYP can be achieved through the competitive coexistence of parallel/sequential mechanisms via pR1→pB/pR1→pR2 transitions,in which the ground state recovery is controlled by a serial of slow reactions of volume-conserving bicycle pedal/hula twist and one-bond flip isomerization with high barrier,affiliating with the fast protonation/deprotonation processes,and hydrophobic-hydrophilic state transformation.A barrierless excited-state(S1)proton transfer,which is exclusively driven by the Asp148 residue introduced in the S65T/H148D GFP mutant,is responsible for the ultrafast formation of the anionic fluorescent state,which can be deactivated by a concerted asynchronous hula-twist photoisomerization.This causes the lower fluorescence quantum yield in S65T/H148D compared to wild-type GFP.Hydrogen-out-of-plane motion plays an important role in the deactivation of the S65T/H148D fluorescent state.