Color Tuning Mechanism Research Articles

Color Tuning in Rhodopsins: The Mechanism for the Spectral Shift between Bacteriorhodopsin and Sensory Rhodopsin II

The mechanism of color tuning in the rhodopsin family of proteins has been studied by comparing the optical properties of the light-driven proton pump bacteriorhodopsin (bR) and the light detector sensory rhodopsin II (sRII). Despite a high structural similarity, the maximal absorption is blue-shifted from 568 nm in bR to 497 nm in sRII. The molecular mechanism of this shift is still a matter of debate, and its clarification sheds light onto the general mechanisms of color tuning in retinal proteins. The calculations employ a combined quantum mechanical/molecular mechanical (QM/MM) technique, using a DFT-based method for ground state properties and the semiempirical OM2/MRCI method and ab initio SORCI method for excited state calculations. The high efficiency of the methodology has allowed us to study a wide variety of aspects including dynamical effects. The absorption shift as well as various mutation experiments and vibrational properties have been successfully reproduced. Our results indicate that several sources contribute to the spectral shift between bR and sRII. The main factors are the counterion region at the extracellular side of retinal and the amino acid composition of the binding pocket. Our analysis allows a distinction and identification of the different effects in detail and leads to a clear picture of the mechanism of color tuning, which is in good agreement with available experimental data.

Electrostatic potential at the retinal of three archaeal rhodopsins: Implications for their different absorption spectra

The color tuning mechanism of the rhodopsin protein family has been in the focus of research for decades. However, the structural basis of the tuning mechanism in general and of the absorption shift between rhodopsins in particular remains under discussion. It is clear that a major determinant for spectral shifts between different rhodopsins are electrostatic interactions between the chromophore retinal and the protein. Based on the Poisson-Boltzmann equation, we computed and compared the electrostatic potential at the retinal of three archaeal rhodopsins: bacteriorhodopsin (BR), halorhodopsin (HR), and sensory rhodopsin II (SRII) for which high-resolution structures are available. These proteins are an excellent test case for understanding the spectral tuning of retinal. The absorption maxima of BR and HR are very similar, whereas the spectrum of SRII is considerably blue shifted--despite the structural similarity between these three proteins. In agreement with their absorption maxima, we find that the electrostatic potential is similar in BR and HR, whereas significant differences are seen for SRII. The decomposition of the electrostatic potential into contributions of individual residues, allowed us to identify seven residues that are responsible for the differences in electrostatic potential between the proteins. Three of these residues are located in the retinal binding pocket and have in fact been shown to account for part of the absorption shift between BR and SRII by mutational studies. One residue is located close to the beta-ionone ring of retinal and the remaining three residues are more than 8 A away from the retinal. These residues have not been discussed before, because they are, partly because of their location, no obvious candidates for the spectral shift among BR, HR, and SRII. However, their contribution to the differences in electrostatic potential is evident. The counterion of the Schiff base, which is frequently discussed to be involved in the spectral tuning, does not contribute to the dissimilarities between the electrostatic potentials.

Mechanism of color tuning in retinal protein: SAC-CI and QM/MM study

The SAC-CI and QM/MM methods were applied to calculations of the absorption peaks of the five retinal proteins, bovine rhodopsin (Rh), bacteriorhodopsin (bR) in BR, K, and KL states, and sensory rhodopsin II (sRII). The results nicely agree with the experimental excitation energies and provided an insight into the mechanism of the large blue shifts in Rh and sRII from BR: geometric distortion in Rh and protein electrostatic effect in sRII. These results indicate that the present approach is useful for studying the excitation spectra and the mechanism of the color tuning in the retinal proteins.

Absorption of Schiff-Base Retinal Chromophores in Vacuo

The absorption spectrum of the all-trans retinal chromophore in the protonated Schiff-base form, that is, the biologically relevant form, has been measured in vacuo, and a maximum is found at 610 nm. The absorption of retinal proteins has hitherto been compared to that of protonated retinal in methanol, where the absorption maximum is at 440 nm. In contrast, the new gas-phase absorption data constitute a well-defined reference for spectral tuning in rhodopsins in an environment devoid of charges and dipoles. They replace the misleading comparison with absorption properties in solvents and lay the basis for reconsidering the molecular mechanisms of color tuning in the large family of retinal proteins. Indeed, our measurement directly shows that protein environments in rhodopsins are blue- rather than red shifting the absorption. The absorption of a retinal model chromophore with a neutral Schiff base is also studied. The data explain the significant blue shift that occurs when metharhodopsin I becomes deprotonated as well as the purple-to-blue transition of bacteriorhodopsin upon acidification.

X-ray structure of sensory rhodopsin II at 2.1-A resolution.

Sensory rhodopsins (SRs) belong to a subfamily of heptahelical transmembrane proteins containing a retinal chromophore. These photoreceptors mediate the cascade of vision in animal eyes and phototaxis in archaebacteria and unicellular flagellated algae. Signal transduction by these photoreceptors occurs by means of transducer proteins. The two archaebacterial sensory rhodopsins SRI and SRII are coupled to the membrane-bound HtrI and HtrII transducer proteins. Activation of these proteins initiates phosphorylation cascades that modulate the flagellar motors, resulting in either attractant (SRI) or repellent (SRII) phototaxis. In addition, transducer-free SRI and SRII were shown to operate as proton pumps, analogous to bacteriorhodopsin. Here, we present the x-ray structure of SRII from Natronobacterium pharaonis (pSRII) at 2.1-A resolution, revealing a unique molecular architecture of the retinal-binding pocket. In particular, the structure of pSRII exhibits a largely unbent conformation of the retinal (as compared with bacteriorhodopsin and halorhodopsin), a hydroxyl group of Thr-204 in the vicinity of the Schiff base, and an outward orientation of the guanidinium group of Arg-72. Furthermore, the structure reveals a putative chloride ion that is coupled to the Schiff base by means of a hydrogen-bond network and a unique, positively charged surface patch for a probable interaction with HtrII. The high-resolution structure of pSRII provides a structural basis to elucidate the mechanisms of phototransduction and color tuning.

Color Tuning Mechanism of Human Red and Green Visual Pigments

We investigated the molecular mechanism of a rather large red shift of 31 nm in a human red pigment compared with a human green pigment. In this analysis, we paid special attention to the phenomenon of nonadditivity of spectral shifts due to substitution of the key amino acids (OH-bearing amino acids) and the phenomenon of cooperativity by which the spectral shifts due to substitution of the key amino acids in the protein environment of red pigment are about 1.5 times larger than that in the protein environment of green pigment. The analysis was made by using a model of three active sites on which the key amino acids are located and four effective sites by which the effect of the key amino acids is modified. As a result, we found that the interaction between the active sites that occurs through the repolarization of the chromophore induced by the key amino acid is essential for the nonadditivity phenomenon. We also found that the interaction between the active site and the effective site plays a major role in the cooperativity phenomenon. More directly, we say that the highly polarizable property of the chromophore is the origin of the rather large red shift in red pigment. Based on these analyses, we conclude that the interaction between the polarizable chromophore and the protein moiety has the capability of producing a significant spectral shift, at least 1000 cm-1, even by substitution of moderate polar residues of the OH-bearing amino acids.

Color Tuning Mechanism of Human Red and Green Visual Pigments

We investigated the molecular mechanism of a rather large red shift of 31 nm in a human red pigment compared with a human green pigment. In this analysis, we paid special attention to the phenomenon of nonadditivity of spectral shifts due to substitution of the key amino acids (OH-bearing amino acids) and the phenomenon of cooperativity by which the spectral shifts due to substitution of the key amino acids in the protein environment of red pigment are about 1.5 times larger than that in the protein environment of green pigment. The analysis was made by using a model of three active sites on which the key amino acids are located and four effective sites by which the effect of the key amino acids is modified. As a result, we found that the interaction between the active sites that occurs through the repolarization of the chromophore induced by the key amino acid is essential for the nonadditivity phenomenon. We also found that the interaction between the active site and the effective site plays a major role in the cooperativity phenomenon. More directly, we say that the highly polarizable property of the chromophore is the origin of the rather large red shift in red pigment. Based on these analyses, we conclude that the interaction between the polarizable chromophore and the protein moiety has the capability of producing a significant spectral shift, at least 1000 cm1, even by substitution of moderate polar residues of the OH-bearing amino acids.