Abstract
Abstract: :
Purpose: The human retina contains high amounts of the xanthophyll carotenoids lutein [(3R,3'R, 6' R)-ß,ϵ-carotene-3,3' diol] and zeaxanthin [a mixture of (3R,3'R)-ß, ß-carotene-3,3'diol and (3R,3'S-meso)-ß,ß-carotene-3,3' diol]. Their uptake and stabilization is thought to be mediated by specific xanthophyll binding proteins (XBP). We have examined the interactions of XBP with its carotenoid ligands by circular dichroism (CD) spectroscopy. Method: Xanthophyll-protein interactions were studied for their chiral characteristics using CD on partially purified detergent-soluble preparations of XBP from human peripheral retinas. Exogenous carotenoid ligands were added to XBP in tetrahydrofuran (THF) and incubated overnight. Unbound ligands were extracted into hexane. All CD experiments were performed at 4°C using suitable controls. Results: 3R,3'S-meso-zeaxanthin is an optically inactive xanthophyll, and therefore it gives no CD spectrum in organic solvents such as THF, unlike the distinct CD spectra obtained from optically active lutein and 3R,3'R zeaxanthin. We found that 3R,3'S-meso-zeaxanthin exhibited a strong induced CD spectrum in association with XBP. Lutein and zeaxanthin also displayed slight alterations in their CD spectra in association with XBP. Conclusion: These results indicate that CD spectroscopy is an excellent method to probe specific interactions of XBP with its carotenoid ligands. These findings indicate that 3R,3'S-meso-zeaxanthin when bound to XBP is likely to be held in a rigid binding pocket on the protein.
Keywords: 308 age-related macular degeneration • 337 carotenoids/carotenoid binding proteins