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Unbiased simulations reveal the inward-facing conformation of the human serotonin transporter and Na(+) ion release.

ABSTRACT: Monoamine transporters are responsible for termination of synaptic signaling and are involved in depression, control of appetite, and anxiety amongst other neurological processes. Despite extensive efforts, the structures of the monoamine transporters and the transport mechanism of ions and substrates are still largely unknown. Structural knowledge of the human serotonin transporter (hSERT) is much awaited for understanding the mechanistic details of substrate translocation and binding of antidepressants and drugs of abuse. The publication of the crystal structure of the homologous leucine transporter has resulted in homology models of the monoamine transporters. Here we present extended molecular dynamics simulations of an experimentally supported homology model of hSERT with and without the natural substrate yielding a total of more than 1.5 µs of simulation of the protein dimer. The simulations reveal a transition of hSERT from an outward-facing occluded conformation to an inward-facing conformation in a one-substrate-bound state. Simulations with a second substrate in the proposed symport effector site did not lead to conformational changes associated with translocation. The central substrate binding site becomes fully exposed to the cytoplasm leaving both the Na(+)-ion in the Na2-site and the substrate in direct contact with the cytoplasm through water interactions. The simulations reveal how sodium is released and show indications of early events of substrate transport. The notion that ion dissociation from the Na2-site drives translocation is supported by experimental studies of a Na2-site mutant. Transmembrane helices (TMs) 1 and 6 are identified as the helices involved in the largest movements during transport.

SUBMITTER: Koldso H

PROVIDER: S-EPMC3203053 | biostudies-literature | 2011 Oct

REPOSITORIES: biostudies-literature

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Unbiased simulations reveal the inward-facing conformation of the human serotonin transporter and Na(+) ion release.

Koldsø Heidi H Noer Pernille P Grouleff Julie J Autzen Henriette Elisabeth HE Sinning Steffen S Schiøtt Birgit B

PLoS computational biology 20111027 10

Monoamine transporters are responsible for termination of synaptic signaling and are involved in depression, control of appetite, and anxiety amongst other neurological processes. Despite extensive efforts, the structures of the monoamine transporters and the transport mechanism of ions and substrates are still largely unknown. Structural knowledge of the human serotonin transporter (hSERT) is much awaited for understanding the mechanistic details of substrate translocation and binding of antide ...[more]

PMID: 22046120

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Project description:Employing molecular dynamics (MD) simulations, the pathway and mechanism of substrate unbinding from the inward-facing state of the Na(+)-coupled galactose transporter, vSGLT, have been investigated. During a 200-ns equilibrium simulation, repeated spontaneous unbinding events of the substrate from its binding site have been observed. In contrast to the previously proposed gating role of a tyrosine residue (Y263), the unbinding mechanism captured in the present equilibrium simulation does not rely on the displacement and/or rotation of this side chain. Rather, the unbinding involves an initial lateral displacement of the substrate out of the binding site which allows the substrate to completely emerge from the region covered by the side chain of Y263 without any noticeable conformational changes of the latter. Starting with the snapshots taken from this equilibrium simulation with the substrate outside the binding site, steered MD (SMD) simulations were then used to probe the translocation of the substrate along the remaining of the release pathway within the protein's lumen and to characterize the nature of protein-substrate interactions involved in the process. Combining the results of the equilibrium and SMD simulations, we provide a description of the full translocation pathway for the substrate release from the binding site into the cytoplasm. Residues E68, N142, T431, and N267 facilitate the initial substrate's displacement out of the binding site, while the translocation of the substrate along the remainder of the exit pathway formed between TM6 and TM8 is facilitated by H-bond interactions between the substrate and a series of conserved, polar residues (Y138, N267, R273, S365, S368, N371, S372, and T375). The observed molecular events indicate that no gating is required for the release of the substrate from the crystallographically captured structure of the inward-facing state of SGLT, suggesting that this conformation might represent an open, rather than occluded, state of the transporter. This article is part of a Special Issue entitled: Membrane protein structure and function.

Dataset Information

Unbiased simulations reveal the inward-facing conformation of the human serotonin transporter and Na(+) ion release.

Publications

Unbiased simulations reveal the inward-facing conformation of the human serotonin transporter and Na(+) ion release.

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