A Hybrid Methods Approach to the Structural Determination of Photoactivation in the Orange Carotenoid Protein

Abstract

Photoprotective mechanisms are of fundamental importance for the survival of photosynthetic organisms. In cyanobacteria, the orange carotenoid protein (OCP), when activated by intense blue light, binds to the light-harvesting antenna and triggers the dissipation of excess captured light energy. Using a combination of small angle X-ray scattering (SAXS), X-ray hydroxyl radical footprinting, circular di- chroism, and H/D exchange mass spectrometry, we identified both the local and global structural changes in the OCP upon photo- activation. SAXS and H/D exchange data showed that global tertiary structural changes, including complete domain dissociation, occur upon photoactivation, but with alteration of secondary structure confined to only the N terminus of the OCP. High flux radiolytic labeling identified confrontational dynamics of the H-bonding network associated with conserved residues and structural water molecules. Collectively, these data provide experimental evidence for an ensemble of local and global structural changes, upon activation of the OCP, that are essential for photoprotection.