Chemical and structural investigation of the paroxetine-human serotonin transporter complex
- Oregon Health and Science University, United States
- 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.
Article and author information
Author details
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.
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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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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