title: Conversion of a light-driven proton pump into a light-gated ion channel.
abstract: Interest in microbial rhodopsins with ion pumping activity has been revitalized in the context of optogenetics, where light-driven ion pumps are used for cell hyperpolarization and voltage sensing. We identified an opsin-encoding gene (CsR) in the genome of the arctic alga Coccomyxa subellipsoidea C-169 that can produce large photocurrents in Xenopus oocytes. We used this property to analyze the function of individual residues in proton pumping. Modification of the highly conserved proton shuttling residue R83 or its interaction partner Y57 strongly reduced pumping power. Moreover, this mutation converted CsR at moderate electrochemical load into an operational proton channel with inward or outward rectification depending on the amino acid substitution. Together with molecular dynamics simulations, these data demonstrate that CsR-R83 and its interacting partner Y57 in conjunction with water molecules forms a proton shuttle that blocks passive proton flux during the dark-state but promotes proton movement uphill upon illumination.
authors.affiliation: Institute of Biology, Experimental Biophysics, Humboldt-Universität zu Berlin, 10115 Berlin, Germany.
authors.forename: A
authors.initials: A
authors.lastname: Vogt
authors.affiliation: Institute of Physical Chemistry, Karlsruhe Institute of Technology, 76131 Karlsruhe, Germany.
authors.forename: Y
authors.initials: Y
authors.lastname: Guo
authors.affiliation: Institute of Biology, Experimental Biophysics, Humboldt-Universität zu Berlin, 10115 Berlin, Germany.
authors.forename: S P
authors.initials: SP
authors.lastname: Tsunoda
authors.affiliation: Department of Biochemistry, University of Delhi South Campus, New Delhi, India.
authors.forename: S
authors.initials: S
authors.lastname: Kateriya
authors.affiliation: Institute of Physical Chemistry, Karlsruhe Institute of Technology, 76131 Karlsruhe, Germany.
authors.forename: M
authors.initials: M
authors.lastname: Elstner
authors.affiliation: Institute of Biology, Experimental Biophysics, Humboldt-Universität zu Berlin, 10115 Berlin, Germany.
authors.forename: P
authors.initials: P
authors.lastname: Hegemann
mesh_terms: D000818:Animals; D000460:Chlorophyta; D057809:HEK293 Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008564:Membrane Potentials; D056004:Molecular Dynamics Simulation; D009865:Oocytes / Q000378:metabolism; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics*; D015202:Protein Engineering; D017494:Proton Pumps / Q000737:chemistry / Q000235:genetics*; D012243:Rhodopsin / Q000737:chemistry / Q000235:genetics*; D014982:Xenopus laevis
chemical_list: D010940:Plant Proteins; D017494:Proton Pumps; D012243:Rhodopsin