Binding and Orientation of Tricyclic Antidepressants within the Central Substrate Site of the Human Serotonin Transporter

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  • Centre for Psychiatric Research
  • Section for Anthropology and Ethnography
  • iNano-School
  • Department of Chemistry
  • Department of Molecular Biology
  • Interdisciplinary Nanoscience Center
Tricyclic antidepressants (TCA) have been used for decades but their orientation and molecular interactions with their primary target is yet unsettled. The recent finding of a TCA binding site in the cytoplasmatic vestibule of a bacterial leucine transporter, LeuT, 11 Angstrom above the central site has prompted debate about whether this vestibular site in the bacterial transporter is relevant for antidepressant binding to their relevant physiological target, the human serotonin transporter (hSERT). We present an experimentally validated structural model of imipramine and analogous TCAs in the central substrate binding site of the human serotonin transporter (hSERT). Two possible binding modes were observed from induced fit docking (IFD) calculations. We experimentally validated a single binding mode by combining mutagenesis of hSERT with uptake inhibition studies of different TCA analogs according to the Paired Mutation Ligand Analog Complementation (PaMLAC) paradigm. Using this experimental method we identify a salt bridge between the tertiary aliphatic amine and Asp98. Furthermore, the 7-position of the imipramine ring is found vicinal to Phe335 and the hydrophobic pocket lined by Ala173 and Thr439 is utilized by 3-substituents. These protein-ligand contact points unambigously orientate the TCA within the central binding site and reveals differences between substrate binding and inhibitor binding giving important clues to the inhibition mechanism. Consonant with the well established competitive inhibition of uptake by TCAs, the resulting binding site for TCAs in hSERT is fully overlapping with the serotonin binding site in hSERT and dissimilar to the low affinity noncompetitive TCA site reported in LeuT.
Original languageEnglish
JournalJournal of Biological Chemistry
Pages (from-to)8363-8374
Number of pages12
Publication statusPublished - 2010

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