Abstract
<jats:p>We present here a synergistic experimental/modeling study of the role of water molecules in Orange Carotenoid Protein (OCP). Molecular dynamics (MD) simulations were performed for OCP in its resting orange state with three different chromophores: canthaxanthin (CAN), echinenone (ECH), and 3-hydroxyechinenone (HCN). Whereas all three ketocarotenoids have a C=O moiety on of their β1-ring which is involved in two hydrogen bonds with Trp288 and Tyr201 residues on the C-Terminal Domain, the situation appears very different on β2-ring side in the N-Terminal Domain (NTD), where the interaction with the environment is different in three cases. This is not surprising, since ECH has no substituent in the β2-ring capable of getting involved in hydrogen bond interaction with the NTD donor/acceptor groups, whereas HCN and CAN have. In particular, MD simulations show that OCP-CAN is involved in specific H-bond interactions with two water molecules inside the NTD. HCN, the other carotenoid bearing a group potentially involved in hydrogen bond(s) in the β2ring, does not show such specific and long-living interaction with water molecules, but rather weak and fast exchanging H-bonds. Interestingly, UV-Vis spectroscopy shows that only OCP-CAN is self-activated to the red state upon dehydration. Furthermore, time-resolved FTIR difference experiments suggest that the red form produced upon dehydration is very similar to the red form observed upon photoactivation. Altogether, these results show that in OCP-CAN specific water molecules play a key role in stabilizing carotenoid configuration in the orange state, and suggest that upon dehydration these water molecules do not stabilize the orange form any more, possibly because of their depletion. As a consequence, the red state is produced.</jats:p>