Resonance Raman spectroscopy and quantum chemical modeling studies of protein-astaxanthin interactions in alpha-crustacyanin (major blue carotenoprotein complex in carapace of lobster, Homarus gammarus)

Resonance Raman spectroscopy and quantum chemical calculations were used to investigate the molecular origin of the large redshift assumed by the elec tronic absorption spectrum of astaxanthin in alpha-crustacyanin, the major blue carotenoprotein from the carapace of the lobster, Homarus gammarus. Re sonance Raman spectra of alpha-crustacyanin reconstituted with specifically C-13-labeled astaxanthins at the positions 15, 15,15', 14,14', 13,13', 12, 12', or 20,20' were recorded. This approach enabled us to obtain informatio n about the effect of the ligand-protein interactions an the geometry of th e astaxanthin chromophore in the ground electronic state. The magnitude of the downshifts of the C=C stretching modes for each labeled compound indica te that the main perturbation on the central part of the polyene chain is n ot homogeneous. In addition, changes in the 1250-1400 cm(-1) spectral range indicate that the geometry of the astaxanthin polyene chain is moderately changed upon binding to the protein. Semiempirical quantum chemical modelin g studies (Austin method 1) show that the geometry change cannot be solely responsible for the bathochromic shift from 480 to 632 nm of protein-bound astaxanthin. The calculations are consistent with a polarization mechanism that involves the protonation or another interaction with a positive ionic species of comparable magnitude with both ketofunctionalities of the astaxa nthin-chromophore and support the changes observed in the resonance Raman a nd visible absorption spectra. The results are in good agreement with the c onclusions that were drawn on the basis of a study of the charge densities in the chromophore in alpha-crustacyanin by solid-state NMR spectroscopy. F rom the results the dramatic bathochromic skiff can be explained not only f rom a change in the ground electronic state conformation but also from an i nteraction in the excited electronic state that significantly decreases the energy of the pi-antibonding C=O orbitals and the HOMO-LUMO gap. (C) 1999 John Wiley & Sons, Inc.