PROTEIN-CHROMOPHORE INTERACTIONS IN ALPHA-CRUSTACYANIN, THE MAJOR BLUE CAROTENOPROTEIN FROM THE CARAPACE OF THE LOBSTER, HOMARUS-GAMMARUS - A STUDY BY C-13 MAGIC-ANGLE-SPINNING NMR

MAS (magic angle spinning) C-13 MMR has been used to study protein-chromophore interactions in alpha-crustacyanin, the blue astaxanthin-binding carotenoprotein of the lobster, Homarus gammarus, reconstituted with astaxanthins labelled with C-13 at the 14,14' or 15,15' positions. Two signals are seen for alpha-crustacyanin containing [14,14'-C-13(2)]astaxanthin, shifted 6.9 and 4.0 ppm downfield from the 134.1 ppm signal of uncomplexed astaxanthin in the solid state. With alpha-crustacyanin containing [15,15'-C-13(2)]astaxanthin, one essentially unshifted broad signal is seen. Hence binding to the protein causes a decrease in electronic charge density, providing the first experimental evidence that a charge redistribution mechanism contributes to the bathochromic shift of the astaxanthin in alpha-crustacyanin, in agreement with inferences based on resonance Raman data [Salares, et al. (1979) Biochim. Biophys. Acta 576, 176-191]. The splitting of the 14 and 14' signals provides evidence for asymmetric binding of each astaxanthin molecule by the protein.