Chemical and structural investigation of the paroxetine-human serotonin transporter complex

  1. Jonathan A Coleman
  2. Vikas Navratna
  3. Daniele Antermite
  4. Dongxue Yang
  5. James A Bull
  6. Eric Gouaux  Is a corresponding author
  1. Oregon Health and Science University, United States
  2. Imperial College London, United Kingdom

Abstract

Antidepressants target the serotonin transporter (SERT) by inhibiting serotonin reuptake. Structural and biochemical studies aiming to understand binding of small-molecules to conformationally dynamic transporters like SERT often require thermostabilizing mutations and antibodies to stabilize a specific conformation, leading to questions about relationships of these structures to the bonafide conformation and inhibitor binding poses of wild-type transporter. To address these concerns, we determined the structures of ∆N72/∆C13 and ts2-inactive SERT bound to paroxetine analogues using single-particle cryo-EM and x-ray crystallography, respectively. We synthesized enantiopure analogues of paroxetine containing either bromine or iodine instead of fluorine. We exploited the anomalous scattering of bromine and iodine to define the pose of these inhibitors and investigated inhibitor binding to Asn177 mutants of ts2-active SERT. These studies provide mutually consistent insights into how paroxetine and its analogues bind to the central substrate-binding site of SERT, stabilize the outward-open conformation, and inhibit serotonin transport.

Data availability

The coordinates and associated volumes for the cryo-EM reconstruction of SERT 8B6 Fab paroxetine, Br-paroxetine, and I-paroxetine datasets have been deposited in the PDB (https://www.rcsb.org/) and Electron Microscopy Data Bank (EMDB; https://www.ebi.ac.uk/pdbe/emdb/) under the accession codes 6VRH and 21368, 6VRK and 21369, and 6VRL and 21370, respectively. The half maps for each dataset have also been deposited in the EMDB (https://www.ebi.ac.uk/pdbe/emdb/). The x-ray coordinates for Br-paroxetine and I-paroxetine have been deposited in the PDB (https://www.rcsb.org/) under accession codes 6W2B and 6W2C, respectively.

The following data sets were generated

Article and author information

Author details

  1. Jonathan A Coleman

    Vollum Institute, Oregon Health and Science University, Portland, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Vikas Navratna

    Vollum Institute, Oregon Health and Science University, Portland, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Daniele Antermite

    Chemistry, Imperial College London, London, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
  4. Dongxue Yang

    Vollum Institute, Oregon Health and Science University, Portland, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. James A Bull

    Chemistry, Imperial College London, London, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-3993-5818
  6. Eric Gouaux

    Vollum Institute, Oregon Health and Science University, Portland, United States
    For correspondence
    gouauxe@ohsu.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-8549-2360

Funding

National Institutes of Health (5R37MH070039)

  • Eric Gouaux

Howard Hughes Medical Institute (N/A)

  • Eric Gouaux

Royal Society (UF140161)

  • James A Bull

Royal Society (RG150444)

  • James A Bull

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

Reviewing Editor

  1. Lucy R Forrest, National Institute of Neurological Disorders and Stroke, National Institutes of Health, United States

Publication history

  1. Received: February 27, 2020
  2. Accepted: July 2, 2020
  3. Accepted Manuscript published: July 3, 2020 (version 1)
  4. Version of Record published: September 3, 2020 (version 2)

Copyright

© 2020, Coleman 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. Jonathan A Coleman
  2. Vikas Navratna
  3. Daniele Antermite
  4. Dongxue Yang
  5. James A Bull
  6. Eric Gouaux
(2020)
Chemical and structural investigation of the paroxetine-human serotonin transporter complex
eLife 9:e56427.
https://doi.org/10.7554/eLife.56427

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