Genetically encoded photocrosslinkers locate the high-affinity binding site of antidepressant drugs in the human serotonin transporter

Hafsteinn Rannversson; Jacob Andersen; Lena Sørensen Hall; Benny Bang-Andersen; Minyoung Park; Thomas Huber; Thomas P Sakmar; Kristian Strømgaard

doi:10.1038/ncomms11261

Genetically encoded photocrosslinkers locate the high-affinity binding site of antidepressant drugs in the human serotonin transporter

Hafsteinn Rannversson, Jacob Andersen, Lena Sørensen Hall, Benny Bang-Andersen, Minyoung Park, Thomas Huber, Thomas P Sakmar, Kristian Strømgaard

34 Citationer (Scopus)

Abstract

Despite the well-established role of the human serotonin transporter (hSERT) in the treatment of depression, the molecular details of antidepressant drug binding are still not fully understood. Here we utilize amber codon suppression in a membrane-bound transporter protein to encode photocrosslinking unnatural amino acids (UAAs) into 75 different positions in hSERT. UAAs are incorporated with high specificity, and functionally active transporters have similar transport properties and pharmacological profiles compared with wild-type transporters. We employ ultraviolet-induced crosslinking with p-azido-L-phenylalanine (azF) at selected positions in hSERT to map the binding site of imipramine, a prototypical tricyclic antidepressant, and vortioxetine, a novel multimodal antidepressant. We find that the two antidepressants crosslink with azF incorporated at different positions within the central substrate-binding site of hSERT, while no crosslinking is observed at the vestibular-binding site. Taken together, our data provide direct evidence for defining the high-affinity antidepressant binding site in hSERT.

Originalsprog	Engelsk
Artikelnummer	11261
Tidsskrift	Nature Communications
Vol/bind	7
Sider (fra-til)	1-9
Antal sider	9
ISSN	2041-1723
DOI	https://doi.org/10.1038/ncomms11261
Status	Udgivet - 19 apr. 2016

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10.1038/ncomms11261

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title = "Genetically encoded photocrosslinkers locate the high-affinity binding site of antidepressant drugs in the human serotonin transporter",

abstract = "Despite the well-established role of the human serotonin transporter (hSERT) in the treatment of depression, the molecular details of antidepressant drug binding are still not fully understood. Here we utilize amber codon suppression in a membrane-bound transporter protein to encode photocrosslinking unnatural amino acids (UAAs) into 75 different positions in hSERT. UAAs are incorporated with high specificity, and functionally active transporters have similar transport properties and pharmacological profiles compared with wild-type transporters. We employ ultraviolet-induced crosslinking with p-azido-L-phenylalanine (azF) at selected positions in hSERT to map the binding site of imipramine, a prototypical tricyclic antidepressant, and vortioxetine, a novel multimodal antidepressant. We find that the two antidepressants crosslink with azF incorporated at different positions within the central substrate-binding site of hSERT, while no crosslinking is observed at the vestibular-binding site. Taken together, our data provide direct evidence for defining the high-affinity antidepressant binding site in hSERT.",

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author = "Hafsteinn Rannversson and Jacob Andersen and Hall, {Lena S{\o}rensen} and Benny Bang-Andersen and Minyoung Park and Thomas Huber and Sakmar, {Thomas P} and Kristian Str{\o}mgaard",

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AU - Rannversson, Hafsteinn

AU - Andersen, Jacob

AU - Hall, Lena Sørensen

AU - Bang-Andersen, Benny

AU - Park, Minyoung

AU - Huber, Thomas

AU - Sakmar, Thomas P

AU - Strømgaard, Kristian

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N2 - Despite the well-established role of the human serotonin transporter (hSERT) in the treatment of depression, the molecular details of antidepressant drug binding are still not fully understood. Here we utilize amber codon suppression in a membrane-bound transporter protein to encode photocrosslinking unnatural amino acids (UAAs) into 75 different positions in hSERT. UAAs are incorporated with high specificity, and functionally active transporters have similar transport properties and pharmacological profiles compared with wild-type transporters. We employ ultraviolet-induced crosslinking with p-azido-L-phenylalanine (azF) at selected positions in hSERT to map the binding site of imipramine, a prototypical tricyclic antidepressant, and vortioxetine, a novel multimodal antidepressant. We find that the two antidepressants crosslink with azF incorporated at different positions within the central substrate-binding site of hSERT, while no crosslinking is observed at the vestibular-binding site. Taken together, our data provide direct evidence for defining the high-affinity antidepressant binding site in hSERT.

AB - Despite the well-established role of the human serotonin transporter (hSERT) in the treatment of depression, the molecular details of antidepressant drug binding are still not fully understood. Here we utilize amber codon suppression in a membrane-bound transporter protein to encode photocrosslinking unnatural amino acids (UAAs) into 75 different positions in hSERT. UAAs are incorporated with high specificity, and functionally active transporters have similar transport properties and pharmacological profiles compared with wild-type transporters. We employ ultraviolet-induced crosslinking with p-azido-L-phenylalanine (azF) at selected positions in hSERT to map the binding site of imipramine, a prototypical tricyclic antidepressant, and vortioxetine, a novel multimodal antidepressant. We find that the two antidepressants crosslink with azF incorporated at different positions within the central substrate-binding site of hSERT, while no crosslinking is observed at the vestibular-binding site. Taken together, our data provide direct evidence for defining the high-affinity antidepressant binding site in hSERT.

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KW - Depression

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KW - Models, Molecular

KW - Molecular Structure

KW - Mutation

KW - Phenylalanine

KW - Photochemical Processes

KW - Protein Binding

KW - Protein Structure, Tertiary

KW - Serotonin Plasma Membrane Transport Proteins

KW - Ultraviolet Rays

KW - Journal Article

KW - Research Support, Non-U.S. Gov't

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JO - Nature Communications

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ER -

Genetically encoded photocrosslinkers locate the high-affinity binding site of antidepressant drugs in the human serotonin transporter

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