Takahiro Hirano

Structural Models of the Photointermediates in the Rhodopsin Photocascade, Lumirhodopsin, Metarhodopsin I, and Metarhodopsin II

Model building of the two photointermediates, lumirhodopsin and metarhodopsin I, and the activated form of rhodopsin, metarhodopsin II, is described. An outward swing of the C‐terminal portion of transmembrane segment 3, pivoting on Cys110 at the N‐terminal end of transmembrane segment 3, led to structural models of lumirhodopsin and metarhodopsin I. The conformation of the chromophore in the lumirhodopsin and metarhodopsin I models is controlled by the motion of transmembrane segment 3 and agreed closely with the hydrogen‐bonding states of the protonated Schiff base in lumirhodopsin and metarhodopsin I as deduced from their FTIR and resonance Raman spectra and with the negative and positive CD bands of lumirhodopsin and metarhodopsin I, respectively. The structure of metarhodopsin II was constructed by an outward swing of transmembrane segment 3 and the rigid‐body motion of transmembrane segment 6. The arrangement of the entire transmembrane segment of the metarhodopsin II model closely agreed with the electron paramagnetic resonance spectra of spin‐labeled rhodopsin mutants and provided a structural basis for the protonation of Glu134, which is a key process in transducin activation.

Probing for the Threshold Energy for Visual Transduction: Red‐Shifted Visual Pigment Analogs from 3‐Methoxy‐3‐Dehydroretinal and Related Compounds

Abstract— While azulenic retinal analogs failed to yield a red‐shifted visual pigment analog, the 9‐cis isomers of the push‐pull polyenals 3‐methoxy‐3‐dehydroretinal and 14F‐3‐me‐thoxy‐3‐dehydroretinal yielded iodopsin pigment analogs with absorption maxima at, respectively, 663 and 720 nm. The former gave a relatively stable batho product (700 nm) and was able to activate transducin. A lower activity was observed for the latter. One possible explanation for the combined results is that the excitation energies of these red‐shifted pigments are approaching the threshold energy for visual transduction (although at this time we cannot rigorously exclude a role of the added F‐atom in reducing the transducin activity).

Modelling of photointermediates suggests a mechanism of the flip of the β-ionone moiety of the retinylidene chromophore in the rhodopsin photocascade

Light causes an extremely rapid 11-cis-to-all-trans isomerization of the retinylidene chromophore of rhodopsin. This isomerization leads to bleaching intermediates in the photoactivation cascade. An early photointermediate, bathorhodopsin (Batho), which already contains a photoisomerized all-trans retinylidene chromophore, slowly ( 1 sec) decays by conformational changes to metarhodopsin I (Meta I) through lumirhodopsin (Lumi). The cis-trans photoisomerization of the retinylidene chromophore of rhodopsin occurs within the limited space of opsin, which results in a highly strained conformation of the chromophore. In a photoaffinity labeling experiment, Nakanishi et al. showed that a modified -ionone moiety cross-linked Trp265 on transmembrane segment 6 (TM6) both in the rhodopsin and Batho states, which suggests that the cyclohexenyl moiety remains unchanged in the rhodopsin-to-Batho transition. In the subsequent Batho-to-Lumi transition, the moiety flipped from TM6 towards TM4. The flip of the modified -ionone moiety suggests that TM3 and TM4 rearrange to accommodate the modified -ionone moiety, as schematically shown in Figure 1, while the helix

Constraints of Opsin Structure on the Ligand-binding Site: Studies with Ring-fused Retinals¶

Ring‐fused retinal analogs were designed to examine the hula‐twist mode of the photoisomerization of the 9‐cis retinylidene chromophore. Two 9‐cis retinal analogs, the C11–C13 five‐membered ring–fused and the C12–C14 five‐membered ring–fused retinal derivatives, formed the pigments with opsin. The C11–C13 ring‐fused analog was isomerized to a relaxed all‐trans chromophore (λmax > 400 nm) at even −269°C and the Schiff base was kept protonated at 0°C. The C12–C14 ring‐fused analog was converted photochemically to a bathorhodopsin‐like chromophore (λmax= 583 nm) at −196°C, which was further converted to the deprotonated Schiff base at 0°C. The model‐building study suggested that the analogs do not form pigments in the retinal‐binding site of rhodopsin but form pigments with opsin structures, which have larger binding space generated by the movement of transmembrane helices. The molecular dynamics simulation of the isomerization of the analog chromophores provided a twisted C11–C12 double bond for the C12–C14 ring‐fused analog and all relaxed double bonds with a highly twisted C10–C11 bond for the C11–C13 ring‐fused analog. The structural model of the C11–C13 ring‐fused analog chromophore showed a characteristic flip of the cyclohexenyl moiety toward transmembrane segments 3 and 4. The structural models suggested that hula twist is a primary process for the photoisomerization of the analog chromophores.