Retinal β-Ionone Ring-Salinixanthin Interactions in Xanthorhodopsin: A Study Using Artificial Pigments

Xanthorhodopsin (xR) is a retinal protein that contains, in addition to the retinal chromophore, a carotenoid (salinixanthin) that functions as a light-harvesting antenna [Balashov, S. P., et al. (2005) Science 309, 2061-2064]. The center-center distance between the two polyene chains is 12-13 angstrom, but the distance between the two rings of retinal and salinixanthin is surprisingly small (similar to 5 angstrom) with an angle of similar to 45 degrees [Luecke, H., et al. (2008) Proc. Natl. Acad. Sci. U.S.A. 105, 16561-16565]. We aimed to clarify the role of the beta-ionone ring in the binding of retinal to apo-xR, as well as a possible role that the beta-ionone ring plays in fixation of the salinixanthin 4-keto ring. The binding of native retinal and series of synthetic retinal analogues modified in the beta-ionone ring to apo-xR was monitored by absorption and circular dichroism (CD) spectroscopies. The results indicate that the beta-ionone ring modification significantly affected formation of the retinal-protein covalent bond as well as the pigment absorption and CD spectra. It was observed that several retinal analogues, modified in the retinal beta-ionone ring, did not bind to apo-xR and did not form the pigment. Also, none of these analogues induced the fixation of the salinixanthin 4-keto ring. In addition, we show that the native retinal within its binding site adopts exclusively the 6-s-trans ring-chain conformation.