Energy transfer in rhodopsin, N-retinyl-opsin, and rod outer segments

N-retinyl, the chromophore of bleached and reduced rhodopsin, N-retinyl-opsin, was used as a covalently attached fluorescence probe to examine the structure of N-retinyl-opsin and the rod outer segment. The efficiency of energy transfer from the protein part of N-retinyl-opsin to the chromophore is 12 +/- 5%. It is argued that this implies that the N-retinyl-opsin molecule is asymmetrical. Kropf has estimated the efficiency of energy transfer from the protein to the chromophore in native rhodopsin to be about 50%. This difference of efficiencies seems to imply a large movement of the chromophore away from the tryptophans of the opsin after rhodopsin is bleached. From excitation spectrum measurements, it has been found that light absorbed by the protein of the rod outer segments has more action in sensitizing the fluorescence of the chromophore than does light absorbed by the protein part of pure N-retinyl-opsin. Thus, some other tryptophans or tyrosines in either another N-retinyl-opsin molecule or another protein must be close enough (about 28 A) to the chromophore to transfer energy to it. Measurements of the polarization of the fluorescence of the chromophore suggest, however, that the chromophores of neighboring N-retinyl-opsin molecules are more than 20 A apart. Moreover, these neighboring chromophores do not transfer energy to each other, tending to rule out any clustering of chromophores of different N-retinyl-opsin molecules and suggesting that rhodopsin chromophores do not transfer energy to each other.