Fourier transform infrared spectroscopic investigation of rhodopsin structure and its comparison with bacteriorhodopsin.
FT-IR spectroscopy has been used to investigate the conformation of rhodopsin in bovine rod outer segment membranes, dispersed in aqueous suspension in both 2H2O and H2O. Detailed analysis of the amide I band was made, using second-derivative and deconvolution procedures. The frequency of the major amide I component is consistent with the presence of predominantly alpha-helices within the rhodopsin structure. A spectroscopic change occurs at acidic pH with the membranes in both 2H2O and H2O. The results for the membranes dispersed in H2O at pH 7 were used to estimate a value of 0.67 for w (amide II/amide I intensity ratio in H2O). This value of w gives an estimate of the unexchanged amide protons, in rhodopsin, of 51%. The extent of amide proton exchange at acidic p2H (p2H 5 and 2), in 2H2O was also determined. The conformation of rhodopsin in its unbleached and bleached states was investigated but no significant difference in the secondary structure was observed. A comparison, after second-derivative and deconvolution analysis, of the spectra of rhodopsin with that of bacteriorhodopsin shows that both proteins exhibit a similar number of amide I components. However, with bacteriorhodopsin the amide I band occurs at a higher frequency. Bacteriorhodopsin under similar conditions, in 2H2O, has 20% more unexchanged amide protons than does rhodopsin.
Haris
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PI
Department of Protein and Molecular Biology, University of London, Royal Free Hospital School of Medicine, U.K.
Coke
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M
Chapman
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D
D000818:Animals; D001436:Bacteriorhodopsins; D002417:Cattle; D002462:Cell Membrane / Q000032:analysis; D003903:Deuterium; D005583:Fourier Analysis; D006863:Hydrogen-Ion Concentration; D010777:Photochemistry; D010786:Photoreceptor Cells / Q000032:analysis*; D011487:Protein Conformation; D011522:Protons; D012168:Retinal Pigments; D012243:Rhodopsin; D012374:Rod Cell Outer Segment / Q000032:analysis*; D013055:Spectrophotometry, Infrared; D014867:Water
D011522:Protons; D012168:Retinal Pigments; D014867:Water; D001436:Bacteriorhodopsins; D012243:Rhodopsin; D003903:Deuterium