Mechanisms of Spectral Tuning in Blue Cone Visual Pigments

Abstract

Spectral tuning by visual pigments involves the modulation of the physical properties of the chromophore (11-cis-retinal) by amino acid side chains that compose the chromophore-binding pocket. We identified 12 amino acid residues in the human blue cone pigment that might induce the required green-to-blue opsin shift. The simultaneous substitution of nine of these sites in rhodopsin (M86L, G90S, A117G, E122L, A124T, W265Y, A292S, A295S, and A299C) shifted the absorption maximum from 500 to 438 nm, accounting for 2,830 cm ؊1 , or 80%, of the opsin shift between rhodopsin and the blue cone pigment. Raman spectroscopy of mutant pigments shows that the dielectric character and architecture of the chromophore-binding pocket are specifically altered. An increase in the number of dipolar side chains near the protonated Schiff base of retinal increases the ground-excited state energy gap via long range dipoledipole Coulomb interaction. In addition, the W265Y substitution causes a decrease in solvent polarizability near the chromophore ring structure. Finally, two substitutions on transmembrane helix 3 (A117G and E122L) act in combination with the other substitutions to alter the binding-pocket structure, resulting in stronger interaction of the protonated Schiff base group with the surrounding dipolar groups and the counterion. Taken together, these results identify the amino acid side chains and the underlying physical mechanisms responsible for a majority of the opsin shift in blue visual pigments.At the fundamental level, the ability of the visual system to discriminate wavelengths of electromagnetic radiation is determined by the spectral response of the light-absorbing pigments that reside in the photoreceptor cells of the retina. Trichromatic color vision in humans is mediated by three types of cone cells, each of which contains a specific visual pigment. The blue, green, and red cone pigments absorb maximally at ϳ425, 533, and 560 nm, respectively (1-3). Dim-light (scotopic) vision is mediated through rod photoreceptor cells containing the pigment rhodopsin with its absorption maximum ( max ) at 500 nm (4).All visual pigments share the same basic structure. The protein consists of an ϳ40-kDa polypeptide domain folded into seven transmembrane (TM) 1 helical segments. It is covalently bound to an 11-cis-retinal chromophore at a conserved lysine residue on TM helix 7 via a protonated Schiff base (PSB) bond (5-7). An 11-cis-retinal PSB molecule dissolved in an organic solvent such as ethanol has a max at ϳ440 nm. However, its max can range from ϳ420 to 560 nm in visual pigments due to interactions with residues in the binding site of the protein, which modulate the ground-excited electronic (S 0 -S 1 ) transition energy of the retinal PSB. The difference in the max of the pigment from that of the PSB model compound has been termed the "opsin shift" (8), which is also used to refer to the differences in the max values among pigments. Empirical and theoretical studies have identified several mechanisms by which opsin sh...

Ultraviolet pigments in birds evolved from violet pigments by a single amino acid change

“…However, the mutant pigment at pH 4.4, 4.8, 5.5, 6.2, 6.4, 6.6, 7.2, 7.5, 7.8, 8.6, and 11.4 shows identical absorption spectrum to that at pH 6.6 (results not shown). Thus, the various levels of acid treatment suggest that there is no change in its protonation state that influences the electrostatic environment of the chromophore (26). At present, it is not clear why the absorption spectrum of the mutant pigment is broader than that of the UV pigment.…”

“…The amino acid site 84 is located near the Schiff base nitrogen and E107 (corresponds to E113 of the bovine rhodopsin), counterion of the Schiff base (12,26,30). Using bovine rhodopsin, it has been suggested that one or a few water molecules is located in this region (31)(32)(33)(34).…”

Importance of the Broad Regional Interaction for Spectral Tuning in Natronobacterium pharaonis Phoborhodopsin (Sensory Rhodopsin II)

“…This result implied that there might be other amino acid residues determining the color regulation because it is quite conceivable that the change in the environment around the chromophore is one of the most influential factors regulating color. In fact, Sakmar and co-workers (26,27) succeeded in producing a significant blue shift for rhodopsin (500 nm) to 438 nm by simultaneous substitution of nine sites around the chromophore of rhodopsin, whose shift amounts to 80% of the spectral tuning between rhodopsin and the blue cone pigment. Therefore, it is considered that the max among archaeal rhodopsins is also determined by the difference in the environment around the chromophore.…”

The Green-absorbing Drosophila Rh6 Visual Pigment Contains a Blue-shifting Amino Acid Substitution That Is Conserved in Vertebrates

“…Mutagenesis experiments at this site in a variety of pigments have shown that A292S or S292A mutants demonstrate substantial blue and red spectral shifts, respectively (8,(12)(13)(14)(15)(16). As shown in Fig.…”

“…This amino acid substitution red-shifts the max of the Drosophila Rh1 pigment and reciprocally blue-shifts the max of Rh6 pigment. Interestingly, this site also affects the spectral tuning of vertebrate pigments and corresponds to Ala-292 in bovine rhodopsin (8,(12)(13)(14)(15)(16).…”