Methyl Substituents at the 11 or 12 Position of Retinal Profoundly and Differentially Affect Photochemistry and Signalling Activity of Rhodopsin

The C-11 = C-12 double bond of the retinylidene chromophore of rhodopsin holds a central position in its light-induced photoisomerization and hence the photosensory function of this visual pigment. To probe the local environment of the HC-11 = C-12H element we have prepared the 11-methyl and 12-methyl derivatives of 11-Z retinal and incorporated these into opsin to generate the rhodopsin analogs 11-methyl and 12-methyl rhodopsin. These analog pigments form with much slower kinetics and lower efficiency than the native pigment. The initial photochemistry and the signaling activity of the analog pigments were investigated by UV-vis and FTIR spectroscopy, and by a G protein activation assay. Our data indicate that the ultrafast formation of the first photointermediate is strongly perturbed by the presence of an 11-methyl substituent, but much less by a 12-methyl substituent. These results support the current concept of the mechanism of the primary photoisomerization event in rhodopsin. An important stronghold of this concept is an out-of-plane movement of the C-12H element, which is facilitated by torsion as well as extended positive charge delocalization into the C-10–C-13 segment of the chromophore. We argue that this mechanism is maintained principally with a methyl substituent at C-12.