1. Structural Biology and Molecular Biophysics

Cryo-EM structures and functional characterization of the murine lipid scramblase TMEM16F

  1. Carolina Alvadia
  2. Novandy K Lim
  3. Vanessa Clerico Mosina
  4. Gert T Oostergetel
  5. Raimund Dutzler  Is a corresponding author
  6. Cristina Paulino  Is a corresponding author
  1. University of Zürich, Switzerland
  2. University of Groningen, Netherlands
https://doi.org/10.7554/eLife.44365
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  1. Carolina Alvadia
  2. Novandy K Lim
  3. Vanessa Clerico Mosina
  4. Gert T Oostergetel
  5. Raimund Dutzler
  6. Cristina Paulino
(2019)
Cryo-EM structures and functional characterization of the murine lipid scramblase TMEM16F
eLife 8:e44365.
https://doi.org/10.7554/eLife.44365
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Abstract

The lipid scramblase TMEM16F initiates blood coagulation by catalyzing the exposure of phosphatidylserine in platelets. The protein is part of a family of membrane proteins, which encompasses calcium-activated channels for ions and lipids. Here, we reveal features of murine TMEM16F (mTMEM16F) that underlie its function as a lipid scramblase and an ion channel. The cryo-EM data of mTMEM16F in absence and presence of Ca2+ define the ligand-free closed conformation of the protein and the structure of a Ca2+-bound intermediate. Both conformations resemble their counterparts of the scrambling-incompetent anion channel mTMEM16A, yet with distinct differences in the region of ion and lipid permeation. In conjunction with functional data, we demonstrate the relationship between ion conduction and lipid scrambling. Although activated by a common mechanism, both functions appear to be mediated by alternate protein conformations that are at equilibrium in the ligand-bound state.

Data availability

The three-dimensional cryo-EM density maps as well as the modelled coordinated will be deposited in the Electron Microscopy Data Bank and the Protein Data Bank, respectively. The deposition includes the cryo-EM maps, both half-maps, and the mask used for final FSC calculation. The raw data can be provided upon request.

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Author details

  1. Carolina Alvadia

    Department of Biochemistry, University of Zürich, Zürich, Switzerland
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8446-1098

  2. Novandy K Lim

    Department of Biochemistry, University of Zürich, Zürich, Switzerland
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-5098-929X

  3. Vanessa Clerico Mosina

    Department of Structural Biology, University of Groningen, Groningen, Netherlands
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8013-0144

  4. Gert T Oostergetel

    Department of Structural Biology, University of Groningen, Groningen, Netherlands
    Competing interests
    The authors declare that no competing interests exist.

  5. Raimund Dutzler

    Department of Biochemistry, University of Zürich, Zürich, Switzerland
    For correspondence
    dutzler@bioc.uzh.ch
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2193-6129

  6. Cristina Paulino

    Department of Structural Biology, University of Groningen, Groningen, Netherlands
    For correspondence
    c.paulino@rug.nl
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-7017-109X

Funding

Nederlandse Organisatie voor Wetenschappelijk Onderzoek (740.018.016)

  • Cristina Paulino

H2020 European Research Council (339116)

  • Raimund Dutzler

H2020 European Research Council (AnoBest)

  • Raimund Dutzler

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

Copyright

© 2019, Alvadia 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. Carolina Alvadia
  2. Novandy K Lim
  3. Vanessa Clerico Mosina
  4. Gert T Oostergetel
  5. Raimund Dutzler
  6. Cristina Paulino
(2019)
Cryo-EM structures and functional characterization of the murine lipid scramblase TMEM16F
eLife 8:e44365.
https://doi.org/10.7554/eLife.44365

Share this article

https://doi.org/10.7554/eLife.44365

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Further reading

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

    Stepwise activation mechanism of the scramblase nhTMEM16 revealed by cryo-EM

    Valeria Kalienkova, Vanessa Clerico Mosina ... Cristina Paulino
    Research Article Updated

    Scramblases catalyze the movement of lipids between both leaflets of a bilayer. Whereas the X-ray structure of the protein nhTMEM16 has previously revealed the architecture of a Ca2+-dependent lipid scramblase, its regulation mechanism has remained elusive. Here, we have used cryo-electron microscopy and functional assays to address this question. Ca2+-bound and Ca2+-free conformations of nhTMEM16 in detergent and lipid nanodiscs illustrate the interactions with its environment and they reveal the conformational changes underlying its activation. In this process, Ca2+ binding induces a stepwise transition of the catalytic subunit cavity, converting a closed cavity that is shielded from the membrane in the absence of ligand, into a polar furrow that becomes accessible to lipid headgroups in the Ca2+-bound state. Additionally, our structures demonstrate how nhTMEM16 distorts the membrane at both entrances of the subunit cavity, thereby decreasing the energy barrier for lipid movement.

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