A Resonance Raman Study Of the C=C Stretch Modes in Bovine and Octopus Visual Pigments with Isotopically Labeled Retinal Chromophores

Abstract

Previous resonance Raman spectroscopic studies of bovine and octopus rhodopsin and bathorhodopsin in the C-C stretch fingerprint region have shown drastically different spectral patterns, which suggest different chromophore-protein interactions. We have extended our resonance Raman studies of bovine and octopus pigments to the C=C stretch region in order to reveal a more detailed picture about the difference in retinal-protein interactions between these two pigments. The C=C stretch motions of the protonated retinal Schiff base are strongly coupled to form highly delocalized ethylenic modes located in the 1500 to 1650 cm-1 spectral region. In order to decouple these vibrations, a series of 11,12-D2-labeled retinals, with additional 13C labeling at C8, C10, C11 and C14, respectively, are used to determine the difference of specific C=C stretch modes between bovine and octopus pigments. Our results show that the C9=C10 and C13=C14 stretch mode are about 20 cm-1 lower in the Raman spectrum of octopus bathorhodopsin than in bovine bathorhodopsin, while the other C=C stretch modes in these two bathorhodopsins are similar. In contrast, only the C9=C10 stretch mode in octopus rhodopsin is about 10 cm-1 lower than in bovine rhodopsin, while other C=C stretches are similar.

Rhodopsin: Structural Basis of Molecular Physiology

“…A number of experimental approaches have been employed to investigate RET-protein interactions in the membrane-embedded domain of bovine Rho. Several spectroscopic methods such as resonance Raman spectroscopy (98,128,148), Fourier-transform infrared (FTIR)-difference spectroscopy (60), and NMR spectroscopy (52,80,81,251) have been reported. Other approaches have included reconstitution of opsin apoprotein with synthetic retinal analogs (97,154) and photochemical cross-linking (26,170,266).…”

Rhodopsin: Insights from Recent Structural Studies

“…A number of experimental approaches have been employed to investigate RETprotein interactions in the membrane-embedded domain of bovine Rho. Several spectroscopic methods such as resonance Raman spectroscopy (93,117,129), Fourier-transform infrared (FTIR)-difference spectroscopy (51) and NMR spectroscopy (44,74,75,221) have been reported. Other approaches have included reconstitution of opsin apoprotein with synthetic retinal analogues (92,137) and photochemical cross-linking (26,154,232).…”

Energetics and Volume Changes of the Intermediates in the Photolysis of Octopus Rhodopsin at a Physiological Temperature

“…The energy storage mechanism is not understood well yet, but it may come from a combination of the distortion of the retinal polyene chain and the Schiff base linkage, alternation of the electrostatic interaction between the protonated Schiff base and a counterion charge, and changes of the steric interaction between the retinal and the amino acid residues in the binding pocket. Conformational changes of the retinal polyene chain and the Schiff base linkage during the Rh* to Batho step were reported using resonance Raman spectroscopy (Pande et al, 1987;Deng et al, 1991a,b;Huang et al, 1996Huang et al, , 1997. Furthermore, the electrostatic interaction between the protonated Schiff base and a counterion was studied by Fourier transform infrared (FTIR), which revealed that the O-H stretch frequency of a water molecule bound to the Schiff base in Batho was changed from that in Rh (Nishimura et al, 1997).…”

“…These photointermediates have been identified by transient absorption spectroscopy, and they are consecutively called primerhodopsin (Prime), bathorhodopsin (Batho), lumirhodopsin (Lumi), mesorhodopsin (Meso), and acid metarhodopsin (Acid Meta) (Tsuda, 1979;Ohtani et al, 1988;Taiji et al, 1992;Nakagawa et al, 1997). The structural changes of the chromophore, peptide backbone, and water of these intermediates have been studied by resonance Raman (Kitagawa and Tsuda, 1980;Pande et al, 1987;Deng et al, 1991a,b;Huang et al, 1996Huang et al, , 1997Hashimoto et al, 1996), FTIR (Masuda et al, 1993a,b;Bagley et al, 1989;Nishimura et al, 1997), and UV difference absorption spectroscopy (Nakagawa et al, 1997). Recently, we showed that regions of the protein distant from the chromophore are still changing even after the changes in the microenvironment around the chromophore are over.…”