Chimeric microbial rhodopsins for optical activation of Gs-proteins

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Author(s)

    • Yoshida Kazuho
    • Department of Life Science and Applied Chemistry, Nagoya Institute of Technology
    • Sasaki Kengo
    • Department of Life Science and Applied Chemistry, Nagoya Institute of Technology
    • Inoue Keiichi
    • Department of Life Science and Applied Chemistry, Nagoya Institute of Technology|OptoBioTechnology Research Center, Nagoya Institute of Technology|PRESTO, Japan Science and Technology Agency
    • Kandori Hideki
    • Department of Life Science and Applied Chemistry, Nagoya Institute of Technology|OptoBioTechnology Research Center, Nagoya Institute of Technology

Abstract

<p>We previously showed that the chimeric proteins of microbial rhodopsins, such as light-driven proton pump bacteriorhodopsin (BR) and <i>Gloeobacter</i> rhodopsin (GR) that contain cytoplasmic loops of bovine rhodopsin, are able to activate Gt protein upon light absorption. These facts suggest similar protein structural changes in both the light-driven proton pump and animal rhodopsin. Here we report two trials to engineer chimeric rhodopsins, one for the inserted loop, and another for the microbial rhodopsin template. For the former, we successfully activated Gs protein by light through the incorporation of the cytoplasmic loop of β<sub>2</sub>-adrenergic receptor (β<sub>2</sub>AR). For the latter, we did not observe any G-protein activation for the light-driven sodium pump from <i>Indibacter alkaliphilus</i> (<i>Indi</i>R2) or a light-driven chloride pump halorhodopsin from <i>Natronomonas pharaonis</i> (<i>Np</i>HR), whereas the light-driven proton pump GR showed light-dependent G-protein activation. This fact suggests that a helix opening motion is common to G protein coupled receptor (GPCR) and GR, but not to <i>Indi</i>R2 and <i>Np</i>HR. Light-induced difference FTIR spectroscopy revealed similar structural changes between WT and the third loop chimera for each light-driven pump. A helical structural perturbation, which was largest for GR, was further enhanced in the chimera. We conclude that similar structural dynamics that occur on the cytoplasmic side of GPCR are needed to design chimeric microbial rhodopsins.</p>

Journal

  • Biophysics and Physicobiology

    Biophysics and Physicobiology 14(0), 183-190, 2017

    The Biophysical Society of Japan

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