1. Biochemistry and Chemical Biology
  2. Structural Biology and Molecular Biophysics

Crystal structure of a natural light-gated anion channelrhodopsin

  1. Hai Li
  2. Chia-Ying Huang
  3. Elena G Govorunova
  4. Christopher T Schafer
  5. Oleg A Sineshchekov
  6. Meitian Wang
  7. Lei Zheng  Is a corresponding author
  8. John L Spudich  Is a corresponding author
  1. University of Texas Health Science Center at Houston, McGovern Medical School, United States
  2. Swiss Light Source, Paul Scherrer Institute, Switzerland
https://doi.org/10.7554/eLife.41741
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Cite this article
  1. Hai Li
  2. Chia-Ying Huang
  3. Elena G Govorunova
  4. Christopher T Schafer
  5. Oleg A Sineshchekov
  6. Meitian Wang
  7. Lei Zheng
  8. John L Spudich
(2019)
Crystal structure of a natural light-gated anion channelrhodopsin
eLife 8:e41741.
https://doi.org/10.7554/eLife.41741
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Abstract

The anion channelrhodopsin GtACR1 from the alga Guillardia theta is a potent neuron-inhibiting optogenetics tool. Presented here, its X-ray structure at 2.9 Å reveals a tunnel traversing the protein from its extracellular surface to a large cytoplasmic cavity. The tunnel is lined primarily by small polar and aliphatic residues essential for anion conductance. A disulfide-immobilized extracellular cap facilitates channel closing and the ion path is blocked mid-membrane by its photoactive retinylidene chromophore and further by a cytoplasmic side constriction. The structure also reveals a novel photoactive site configuration that maintains the retinylidene Schiff base protonated when the channel is open. These findings suggest a new channelrhodopsin mechanism, in which the Schiff base not only controls gating, but also serves as a direct mediator for anion flux.

Data availability

Diffraction data have been deposited in PDB under the accession code 6EDQ.

The following data sets were generated

Article and author information

Author details

  1. Hai Li

    Department of Biochemistry and Molecular Biology, McGovern Medical School, University of Texas Health Science Center at Houston, McGovern Medical School, Houston, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3969-6709

  2. Chia-Ying Huang

    Macromolecular Crystallography, Swiss Light Source, Paul Scherrer Institute, Villigen, Switzerland
    Competing interests
    No competing interests declared.

  3. Elena G Govorunova

    Center for Membrane Biology, Department of Biochemistry and Molecular Biology, University of Texas Health Science Center at Houston, McGovern Medical School, Houston, United States
    Competing interests
    Elena G Govorunova, and The University of Texas Health Science Center at Houston have filed patent applications that relate to ACRs (PCT application PCT/US2016/023095, entitled Compositions And Methods For Use Of Anion Channel Rhodopsins).

  4. Christopher T Schafer

    Center for Membrane Biology, Department of Biochemistry and Molecular Biology, University of Texas Health Science Center at Houston, McGovern Medical School, Houston, United States
    Competing interests
    No competing interests declared.

  5. Oleg A Sineshchekov

    Center for Membrane Biology, Department of Biochemistry and Molecular Biology, University of Texas Health Science Center at Houston, McGovern Medical School, Houston, United States
    Competing interests
    Oleg A Sineshchekov, and The University of Texas Health Science Center at Houston have filed patent applications that relate to ACRs (PCT application PCT/US2016/023095, entitled Compositions And Methods For Use Of Anion Channel Rhodopsins).

  6. Meitian Wang

    Macromolecular Crystallography, Swiss Light Source, Paul Scherrer Institute, Villigen, Switzerland
    Competing interests
    No competing interests declared.

  7. Lei Zheng

    Center for Membrane Biology, Department of Biochemistry and Molecular Biology, University of Texas Health Science Center at Houston, McGovern Medical School, Houston, United States
    For correspondence
    Lei.Zheng@uth.tmc.edu
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-7789-5234

  8. John L Spudich

    Center for Membrane Biology, Department of Biochemistry and Molecular Biology, University of Texas Health Science Center at Houston, McGovern Medical School, Houston, United States
    For correspondence
    John.L.Spudich@uth.tmc.edu
    Competing interests
    John L Spudich, and The University of Texas Health Science Center at Houston have filed patent applications that relate to ACRs (PCT application PCT/US2016/023095, entitled Compositions And Methods For Use Of Anion Channel Rhodopsins).
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-4167-8590

Funding

National Institutes of Health (R01GM027750)

  • John L Spudich

National Institutes of Health (U01MH109146)

  • John L Spudich

Welch Foundation (AU-0009)

  • John L Spudich

American Heart Association (18TPA34230046)

  • Lei Zheng

Hermann Eye Fund

  • John L Spudich

Marie-Skłodowska-Curie (701647)

  • Chia-Ying Huang

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Sriram Subramaniam, University of British Columbia, Canada

Version history

  1. Received: September 5, 2018
  2. Accepted: January 4, 2019
  3. Accepted Manuscript published: January 7, 2019 (version 1)
  4. Version of Record published: January 17, 2019 (version 2)

Copyright

© 2019, Li et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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  1. Hai Li
  2. Chia-Ying Huang
  3. Elena G Govorunova
  4. Christopher T Schafer
  5. Oleg A Sineshchekov
  6. Meitian Wang
  7. Lei Zheng
  8. John L Spudich
(2019)
Crystal structure of a natural light-gated anion channelrhodopsin
eLife 8:e41741.
https://doi.org/10.7554/eLife.41741

Share this article

https://doi.org/10.7554/eLife.41741

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Further reading

    1. Structural Biology and Molecular Biophysics

    The crystal structure of bromide-bound GtACR1 reveals a pre-activated state in the transmembrane anion tunnel

    Hai Li, Chia-Ying Huang ... John L Spudich
    Research Advance Updated

    The crystal structure of the light-gated anion channel GtACR1 reported in our previous Research Article (Li et al., 2019) revealed a continuous tunnel traversing the protein from extracellular to intracellular pores. We proposed the tunnel as the conductance channel closed by three constrictions: C1 in the extracellular half, mid-membrane C2 containing the photoactive site, and C3 on the cytoplasmic side. Reported here, the crystal structure of bromide-bound GtACR1 reveals structural changes that relax the C1 and C3 constrictions, including a novel salt-bridge switch mechanism involving C1 and the photoactive site. These findings indicate that substrate binding induces a transition from an inactivated state to a pre-activated state in the dark that facilitates channel opening by reducing free energy in the tunnel constrictions. The results provide direct evidence that the tunnel is the closed form of the channel of GtACR1 and shed light on the light-gated channel activation mechanism.

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