Homo sapien Serotonin Transporter (hSERT)

Phu Duong '21 and Allison Hector '21

Contents:

I. Introduction

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    Depression decreases the quality of life for more than 264 million people worldwide [1]. The predominant pathophysiological hypothesis attributes depression to decreased signalling of the monoamine neurotransmitter serotonin (5-HT) at the neural synapse [2]. Serotonin modulates a broad range of brain and bodily functions from sleep and behaviour to body temperature and reproduction. In neurons, serotonin packages into vesicles. When an excitatory signal reaches a presynaptic neuron, the serotonin-packed vesicles translocate to the neuronal junction, releasing the neurotransmitter into the synaptic cleft. Serotonin then binds to serotonin receptors in the postsynaptic neuron activating an excitatory or inhibitory cascade. As long as serotonin remains in the cleft, the signal continues to propagate, until excess serotonin reuptake terminates it [4].


synapse

Fig 1. Illustration of the prototypical interneuron signalling pathway. An excitatory signal reaches the axon terminal, causing release of neurotransmitters into the synaptic cleft. Neurotransmitter ligands outward-facing receptors on the surface of the postsynaptic neuron and thereby induces a molecular or biochemical response. Transporters at the presynaptic terminal remove excess neurotransmitters from the cleft which terminates the signal [4].

ligand

Fig 2. Chemical structures of relevant hSERT ligands. (A) The chemical structure of the monoamine neurotransmitter serotonin (5-HT) and its protonated form, the substrate for hSERT reuptake. (B) Chemical structure of the selective-serotonin reuptake inhibitor (SSRI) antidepressant (S)-citalopram.

   
    Neurons require the Human Serotonin Transporter (hSERT), a member of the neurotransmitter sodium symporter family of transport proteins, to expedite the otherwise slow and energetically unfavorable reuptake of 5-HT [5]. In its outward rectifying conformation bound to sodium and chloride, SERT initiates transport by first binding a second sodium ion, followed by protonated serotonin molecule (5-HT+). Because of the high extracellular sodium concentration maintained by the Na+/K+-ATPase, the bound sodium readily moves down its concentration gradient and into the cell, driving serotonin transport by inducing an inward rectifying conformational change to the protein. Once the protein flips inward to release the second sodium ion and 5-HT+ into the cytoplasm, intracellular potassium binds to SERT, reverting it back to its outward rectifying conformation, and then exits for transport reinitiation (Fig 3) [6].
function

Fig 3. Schematic model of hSERT transport. (1-3) Na+ and Cl- bound hSERT in its outward-open conformation invites binding of second Na+ and endogenous 5-HT+. (4-8) A series of conformational changes ensues until the inward-open conformation is reached at which time a Na+ and 5-HT+ is released into the cytoplasm. (9-12) Intracellular K+ ion binds to the inward-facing transporter, leading to conformational reset of hSERT as it exits the cell [6].

II. General Structure and Function
   
    hSERT possesses twelve transmembrane helices ( ; helix colors correspond to Fig 4) connected by extra- and intracellular loops. TM1-5 and TM6-10, oppositely-oriented structural repeats known as a fold, form the central binding site for ligands and ions. Within this inverted-repeat, helices TM1, TM2, TM6, and TM7 are thought to be the key structural elements that change conformation to carry 5-HT across the membrane, forming or breaking molecular interactions according to gating mechanisms found at the intra- and extracellular vestibules [5]. TM

Fig 4. Representation of the transmembrane topology of the human SERT. Intra- and extracellular loops connect the twelve membrane-spanning helices. The orange (TM1-5) and blue (TM6-10) triangles highlight the inverted structural repeats known as the LeuT structural fold [6].

III. Endogenous Substrate Binding Sites
 
 Ion binding sites. As aforementioned in the Introduction, SERT function is sodium and chloride-dependent. The first binding site, formed by TM1, TM6, and TM7, binds Na+ via ion-dipole interactions with residue backbone and side chains. The ion binding (not depicted) is coordinated by residues from TM2, TM6, and TM7. The crucial second ion, similar to the first Na+, forms ion-dipole interactions with the backbone and side chains of residues from TM1 and TM8 [6][3].

   Serotonin binding at orthosteric site. As of December 2019, there are no X-ray structures of hSERT bound to serotonin. Most recent docking experiments, however, point to several potential residues that could interact with (Fig 2A) based on what is known about the conserved LeuT structural fold. Asp98 selectively hydrogen bonds with the protonated amine of 5-HT+. Ala173 contacts aliphatic regions with hydrophobic interactions, while the aromatic side chains of Phe341 cradles the indole ring. Lastly, Ser439 engages in H-bonding with the hydroxyl of the neurotransmitter [3].

IV. Antidepressant Binding Sites


    Selective-Serotonin Reuptake Inhibitors (SSRIs) are drugs designed to bind to SERT with higher affinity than serotonin, competitively-inhibiting serotonin reuptake and prolonging serotonergic signalling in order to decrease depressive symptoms in patients [5].

    The antidepressant (Fig 2B) occupies both the central and allosteric site of the hSERT. Antidepressant binding at the central site sterically inhibits serotonin binding, while the allosteric-bound antidepressant cooperates to increase and prolong antidepressant-affinity at the central site. In the binding site, the amine of (S)-citalopram, while interacting with the carboxylate of Asp98, also forms cation-pi interaction with Tyr95 which may also with its oxygen. The fluorophenyl is embedded within a of hydrophobic and pi-pi interactions with surrounding aliphatic and aromatic residues, while the cyanophtalane pi-stack with nearby phenylalanines. At the site, the cyanopthaline and fluorophenyl groups participate in aromatic-specific interactions with Arg104 (cation-pi) and Phe556 (aromatic-aromatic), respectively [3].


VI. References

[1] Depression.” World Health Organization, World Health Organization, 4 Dec. 2019, https://www.who.int/news-room/fact-sheets/detail/depression

[2] Cowen, Philip J, and Michael Browning. “What Has Serotonin to Do with Depression?” World Psychiatry : Official Journal of the World Psychiatric Association (WPA), BlackWell Publishing Ltd, June 2015, https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4471964/.

[3]Coleman, Jonathan A., et al. “X-Ray Structures and Mechanism of the Human Serotonin Transporter.” Nature News, Nature Publishing Group, 6 Apr. 2016, https://www.nature.com/articles/nature17629.

[4] “Action Potentials and Synapses.” Queensland Brain Institute, 9 Nov. 2017, https://qbi.uq.edu.au/brain-basics/brain/brain-physiology/action-potentials-and-synapses.

[5] “IVB1. Serotonin Transporter: A Representative Reuptake Pump.” Serotonin Transporter: A Representative Reuptake Pump, https://web.williams.edu/imput/synapse/pages/IVB1.html.

[6] Hellsberg, Eva, et al. “A Structural Model of the Human Serotonin Transporter in an Outward-Occluded State.” A Structural Model of the Human Serotonin Transporter in an Outward-Occluded State, 2019, doi:10.1101/637009.

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