Biology subjects

Shichida, Y.

Publications and source records attributed to Shichida, Y..

3 recordsLinked to original sources

bioRxiv2025

A key spectral tuning site of UV-sensitive vertebrate non-visual opsin Opn5

Opsins are photoreceptive proteins responsible for visual and non-visual photoreceptions in animals. In general, vertebrates have multiple visual and non-visual opsins whose spectral sensitivities range from the UV to the red region. Among these opsins, Opn5 has been widely identified in vertebrates from fishes to primates and functions as a non-visual opsin in various tissues including the retina and brain. Vertebrate Opn5 has been characterized as a UV-sensitive bistable opsin. Thus, Opn5 provides one of the molecular mechanisms determining the short wavelength limit that vertebrates can detect. In this study, we searched for the amino acid residue responsible for the UV light sensitivity of Opn5. Our mutational analysis revealed that Opn5 acquired visible light sensitivity by the substitution of Lys91 with an amino acid other than arginine or tyrosine residue. In addition, the mutations at Lys91 altered the preferential binding of the retinal isomers in Opn5. Therefore, the conservation of Lys91 among vertebrate Opn5 proteins would be necessary to enable Opn5 to work as the shortest wavelength sensor in various tissues.

biochemistry↗

bioRxiv2021

Creation of photocyclic vertebrate rhodopsin by single amino acid substitution

Opsins are universal photoreceptive proteins in animals and can be classified into three types based on their photoreaction properties. Upon light irradiation, vertebrate rhodopsin forms a metastable active state, which cannot revert back to the original dark state via either photoreaction or thermal reaction. By contrast, after photoreception, most opsins form a stable active state which can photo-convert back to the dark state. Moreover, we recently found a novel type of opsins whose activity is regulated by photocycling. However, the molecular mechanism underlying this diversification of opsins remains unknown. In this study, the molecular property of vertebrate rhodopsin successfully converted to the photocyclic and photoreversible properties by a single mutation at position 188. This revealed that the residue at position 188 contributes to the diversification of photoreaction properties of opsins by the regulation of the recovery from the active state to the original dark state.

biochemistry↗

bioRxiv2017

Single-molecule diffusion-based estimation of GPCR activity

G protein-coupled receptors (GPCRs) are major drug targets and have high potential for drug discovery. The development of a method for measuring the activities of GPCRs is essential for pharmacology and drug screening. However, it is difficult to measure the effects of a drug by monitoring the receptor on the cell surface, and changes in the concentrations of downstream signaling molecules, which depend on signaling pathway selectivity of the receptor, are used as an index of the receptor activity. Here, we show that single-molecule imaging analysis provides an alternative method for assessing ligand effects on GPCR. We monitored the dynamics of the diffusion of metabotropic glutamate receptor 3 (mGluR3), a class C GPCR, under various ligand conditions by using total internal reflection fluorescence microscopy (TIRFM). The single-molecule tracking analysis demonstrates that changes in the average diffusion coefficient of mGluR3 quantitatively reflect the ligand-dependent activity. Then, we reveal that the diffusion of receptor molecules is altered by the common physiological events associated with GPCRs, including G protein binding or accumulation in clathrin-coated pits, by inhibition experiments and dual-color single-molecule imaging analysis. We also confirm the generality of agonist-induced diffusion change in class A and B GPCRs, demonstrating that the diffusion coefficient is a good index for estimating the ligand effects on many GPCRs regardless of the phylogenetic groups, chemical properties of the ligands, and G protein-coupling selectivity.\n\nOne Sentence Summary: Single-molecule imaging for evaluating ligand effects on GPCRs by monitoring the diffusion dynamics on the cell surface.

biophysics↗