Conversion of a light-driven proton pump into a light-gated ion channel

Sci Rep. 2015 Nov 24;5:16450. doi: 10.1038/srep16450.

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Chlorophyta
  • HEK293 Cells
  • Humans
  • Hydrogen-Ion Concentration
  • Membrane Potentials
  • Molecular Dynamics Simulation
  • Oocytes / metabolism
  • Plant Proteins / chemistry
  • Plant Proteins / genetics*
  • Protein Engineering
  • Proton Pumps / chemistry
  • Proton Pumps / genetics*
  • Rhodopsin / chemistry
  • Rhodopsin / genetics*
  • Xenopus laevis

Substances

  • Plant Proteins
  • Proton Pumps
  • Rhodopsin