Orientation of retinal in bacteriorhodopsin as studied by cross-linking using a photosensitive analog of retinal.
The photosensitive m-diazirinophenyl analog of retinal (Fig. 1, II) bound to bacterio-opsin at Lys-216 and regenerated a chromophore with lambda max at 470 nm. Photolysis of the complex at 365 nm resulted in covalent cross-linking of the retinal analog to the bacterio-opsin in greater 30% yield. Investigation of the sites of cross-linking between the 3H-labeled retinal analog and the protein showed the peptide fragment (amino acid residues 190-248) to be the main radioactively labeled product. Stepwise Edman degradation showed Ser-193 and Glu-194 to be the predominant sites of cross-linking. These results show that the chromophore in bacteriorhodopsin is inclined towards helix 6 and towards the exterior of the cell. These data also provide information on the approximate angle that the chromophore makes with the plane of the membrane and they require a modification of the current secondary structure model for bacteriorhodopsin.
D000595:Amino Acid Sequence; D001389:Azirines; D001436:Bacteriorhodopsins; D001665:Binding Sites; D002338:Carotenoids; D003432:Cross-Linking Reagents; D007700:Kinetics; D011485:Protein Binding; D011487:Protein Conformation; D012172:Retinaldehyde / Q000031:analogs & derivatives* / Q000032:analysis*; D014801:Vitamin A / Q000031:analogs & derivatives*
D001389:Azirines; D003432:Cross-Linking Reagents; D014801:Vitamin A; D002338:Carotenoids; D001436:Bacteriorhodopsins; C036294:3-diazirinophenylretinal; D012172:Retinaldehyde