Selective Choice of the Efficient Carotenoid Antenna by a Xanthorhodopsin: Controlling Factors for Binding and Excitation Energy Transfer

Despite extensive research on carotenoids and microbial rhodopsins in aquatic environments, a fundamental understanding of the binding requirements of carotenoids that serve as auxiliary light-harvesting antennas for rhodopsins is still lacking. Our recent discovery of 3-hydroxylated xanthophyll-binding proteorhodopsins and xanthorhodopsins prompted us to investigate the role of keto and hydroxy functional groups in carotenoid binding to rhodopsins and their influence on energy transfer to the retinal chromophore. In this study, we examined the binding of 12 carotenoids to rhodopsin Kin4B8 (a protein of the xanthorhodopsin family, GenBank: OP056329) and assessed the energy transfer between the carotenoid and the retinal chromophore. We found that 3-hydroxylated xanthophylls were the most effective light-harvesting antennas among the carotenoids studied. While 4-ketocarotenoids also bound to the protein, their energy transfer efficiency was significantly reduced. In contrast, the presence of a 4-hydroxy group or the substitution of the β-ionone ring by an ϵ-ionone ring completely prevented binding. Furthermore, mutagenesis studies of Kin4B8 suggest that specific residues play a key role in the selective binding of carotenoids. These findings provide valuable insights into the structural determinants of rhodopsin-carotenoid interactions, which may aid in predicting the recruitment of various carotenoid antennas by retinal proteins.