title: Crystal Structure of the 11-cis Isomer of Pharaonis Halorhodopsin: Structural Constraints on Interconversions among Different Isomeric States.
abstract: Like other microbial rhodopsins, the light driven chloride pump halorhodopsin from Natronomonas pharaonis (pHR) contains a mixture of all-trans/15-anti and 13-cis/15-syn isomers in the dark adapted state. A recent crystallographic study of the reaction states of pHR has shown that reaction states with 13-cis/15-syn retinal occur in the anion pumping cycle that is initiated by excitation of the all-trans isomer. In this study, we investigated interconversions among different isomeric states of pHR in the absence of chloride ions. The illumination of chloride free pHR with red light caused a large blue shift in the absorption maximum of the retinal visible band. During this "red adaptation", the content of the 11-cis isomer increased significantly, while the molar ratio of the 13-cis isomer to the all-trans isomer remained unchanged. The results suggest that the thermally activated interconversion between the 13-cis and the all-trans isomers is very rapid. Diffraction data from red adapted crystals showed that accommodation of the retinal chromophore with the 11-cis/15-syn configuration was achieved without a large change in the retinal binding pocket. The measurement of absorption kinetics under illumination showed that the 11-cis isomer, with a λmax at 565 nm, was generated upon excitation of a red-shifted species (λmax = 625 nm) that was present as a minor component in the dark adapted state. It is possible that this red-shifted species mimics an O-like reaction state with 13-cis/15-syn retinal, which was hypothesized to occur at a late stage of the anion pumping cycle.
authors.affiliation: Department of Physics, Graduate School of Science, Nagoya University , Nagoya 464-8602, Japan.
authors.forename: Siu Kit
authors.initials: SK
authors.lastname: Chan
authors.affiliation: Department of Physics, Graduate School of Science, Nagoya University , Nagoya 464-8602, Japan.
authors.forename: Haruki
authors.initials: H
authors.lastname: Kawaguchi
authors.affiliation: Department of Physics, Graduate School of Science, Nagoya University , Nagoya 464-8602, Japan.
authors.forename: Hiroki
authors.initials: H
authors.lastname: Kubo
authors.affiliation: Department of Physics, Graduate School of Science, Nagoya University , Nagoya 464-8602, Japan.
authors.forename: Midori
authors.initials: M
authors.lastname: Murakami
authors.affiliation: Center for Gene Research, Nagoya University , Nagoya 464-8602, Japan.
authors.forename: Kunio
authors.initials: K
authors.lastname: Ihara
authors.affiliation: Department of Physics, Graduate School of Science, Nagoya University , Nagoya 464-8602, Japan.
authors.forename: Kosuke
authors.initials: K
authors.lastname: Maki
authors.affiliation: Department of Physics, Graduate School of Science, Nagoya University , Nagoya 464-8602, Japan.|RIKEN Harima Branch, 1-1-1, Kouto, Sayo, Hyogo, Japan.
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Kouyama
mesh_terms: D018360:Crystallography, X-Ray; D006216:Halobacteriaceae / Q000737:chemistry* / Q000378:metabolism / Q000528:radiation effects; D025602:Halorhodopsins / Q000737:chemistry* / Q000378:metabolism / Q000528:radiation effects; D007700:Kinetics; D008027:Light; D008958:Models, Molecular; D055668:Photochemical Processes; D011487:Protein Conformation; D013053:Spectrophotometry; D013237:Stereoisomerism
chemical_list: D025602:Halorhodopsins