Activation of Orange Carotenoid Protein Requires a photoisomerization

The orange carotenoid protein (OCP) is a photoactive protein involved in the photoprotection of cyanobacterial antenna complexes. Through two H-bonds in its C-terminal domain, OCP binds a keto-carotenoid (such as canthaxanthin) which is involved in the light-activation of OCP and finally acts as a quencher in the protoprotective mechanism. Absorption of blue light by the resting-state orange OCP (OCPO) causes the translocation of the carotenoid into the N-terminal domain and ultimately the separation of the two domains and forming the active red OCPR state. However, the photochemical mechanism that drives H-bond dissociation and carotenoid translocation is still unknown. Here, we demonstrate that a trans-to-cis photoisomerization of OCP-bound canthaxanthin is a prerequisite for enabling carotenoid translocation. We also show that only the cis photoproduct possesses the spectroscopic features observed in transient absorption spectroscopy. Finally, we explore the role of multiphoton absorption in facilitating the recovery of the all-trans canthaxanthin, which is the isomer found in the structure of OCPR bound to the antenna complexes.