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Y. Shichida, H. Tsukamoto, A. Terakita; Molecular Properties of Rhodopsin Exhibiting Binding Ability to Agonist All–Trans–Retinal . Invest. Ophthalmol. Vis. Sci. 2006;47(13):2840.
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We recently found that amphioxus rhodopsin has binding ability to agonist all–trans–retinal and forms a state that activates G protein , whereas all–trans–retinal does not bind to vertebrate opsin to form active meta–II state. To obtain insight into the difference in agonist binding ability between amphioxus rhodopsin and vertebrate rhodopsin, we have investigated molecular properties of amphioxus rhodopsin by means of spectroscopic and biochemical techniques with the aids of site–directed mutagenesis.
Amphioxus opsin tagged with the rho 1D4 epitope was expressed in HEK 293S cells. Site–directed mutants were prepared as described . Absorption spectra of amphioxus rhodopsin and its mutants were recorded with a conventional spectrophotometer. Their photoreactions were monitored by CCD spectrophotometer.
The opsin of amphioxus rhodopsin can bind 11–cis–retinal to form a photoreceptive pigment that can convert to a red–shifted photoproduct through cis–trans isomerization of the chromophore upon photon absorption. The red–shifted photoproduct is also produced by incubation of opsin with all–trans–retinal and it can activate G protein. The opsin exhibits about 50–fold higher affinity for 11–cis–retinal than for all–trans–retinal, and mutational analyses revealed that Trp–265 situated in helix VI is important for the increase in binding affinity to 11–cis–retinal. Photochemical reactions of amphioxus rhodopsin and its mutants were compared with those of bovine rhodopsin and its mutants and conformational changes of the proteins after photon absorption were found to be different between these pigments.
The amphioxus rhodopsin could be a bridge between the vertebrate rhodopsin and other GPCRs for furthering our understanding of the agonist/antagonist binding and G protein activation mechanism. Reference
 H. Tsukamoto, A. Terakita and Y. Shichida (2005) Proc. Natl. Acad. Sci. USA 102, 6303–6308.
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