Molecular Dynamics Simulations of Na⁺/Cl⁻‐Dependent Neurotransmitter Transporters in a Membrane‐Aqueous System

Abstract

We have performed molecular dynamics simulations of a homology model of the human serotonin transporter (hSERT) in a membrane environment and in complex with either the natural substrate 5-HT or the selective serotonin reuptake inhibitor escitalopram. We have also included a transporter homologue, the Aquifex aeolicus leucine transporter (LeuT), in our study to evaluate the applicability of a simple and computationally attractive membrane system. Fluctuations in LeuT extracted from simulations are in good agreement with crystallographic B factors. Furthermore, key interactions identified in the X-ray structure of LeuT are maintained throughout the simulations indicating that our simple membrane system is suitable for studying the transmembrane protein hSERT in complex with 5-HT or escitalopram. For these transporter complexes, only relatively small fluctuations are observed in the ligand-binding cleft. Specific interactions responsible for ligand recognition, are identified in the hSERT-5HT and hSERT-escitalopram complexes. Our findings are in good agreement with predictions from mutagenesis studies.

Transmembrane Domain 6 of the Human Serotonin Transporter Contributes to an Aqueously Accessible Binding Pocket for Serotonin and the Psychostimulant 3,4-Methylene Dioxymethamphetamine

“…In LeuT Aa , TMs 1, 3, 6, and 8 form the leucine binding pocket. Biochemical analyses and homology modeling of SERT proteins predict a similar binding pocket for 5-HT (32)(33)(34)(35)(36). Consistent with these models, pre-structure studies identified residues in hSERT TMs 1 and 3 that confer high affinity interactions and ligand selectivity to substrates and antagonists.…”

“…Impact of Na ϩ on MTSET Inactivation-The LeuT Aa crystal structure and hSERT homology models derived from it predict that TM6 is involved in the coordination of a sodium ion (27,(32)(33)(34)36). Interestingly, replacement of sodium with NMDG in the MTS incubation buffer resulted in protection of three TM6 Cys mutants and exposure of a single mutant (Fig.…”

“…Because hSERT TM6 is predicted to lie adjacent to and share topology with TM1 (27,(32)(33)(34)(35)(36), we explored potential contributions of TM6 to the interactions of 5-HT, MDMA, and cocaine via an assessment of their ability to protect hSERT C109A Cys mutants from inactivation by MTSET and biotinylation with MTSEA-biotin. Before initiation of protection experiments, we examined the sensitivity of each Cys mutant to MTSET at 0.1, 1, and 10 mM to ensure we were using a concentration of MTSET appropriate for detecting either protection or exposure.…”

Mutational Mapping and Modeling of the Binding Site for (S)-Citalopram in the Human Serotonin Transporter

“…Several LeuT-based models of inhibitor binding to mammalian SLC6 transporters have been reported, including a few addressing antidepressant binding in hSERT (30,31,43,46,54). A model for (S)-citalopram, based on the substrate-bound LeuT structure, was recently presented (43).…”

SLC6 Neurotransmitter Transporters: Structure, Function, and Regulation

“…592 itors (Singh et al, 2007Zhou et al, 2009). It is noteworthy that these LeuT structures have been established as valid templates for molecular modeling of the mammalian NTTs, and LeuT is emerging as an important model protein for biophysical and computational studies addressing the molecular dynamics underlying NTT function and pharmacology (Jørgensen et al, 2007a(Jørgensen et al, , 2008Caplan et al, 2008;Celik et al, 2008a;Noskov and Roux, 2008;Noskov, 2008;Shi et al, 2008;Shaikh and Tajkhorshid, 2010;Zhao et al, 2010a,b).…”