Antidepressant binding site in a bacterial homologue of neurotransmitter transporters

Satinder K. Singh, Atsuko Yamashita, Eric Gouaux

Research output: Contribution to journalArticle

306 Citations (Scopus)

Abstract

Sodium-coupled transporters are ubiquitous pumps that harness pre-existing sodium gradients to catalyse the thermodynamically unfavourable uptake of essential nutrients, neurotransmitters and inorganic ions across the lipid bilayer. Dysfunction of these integral membrane proteins has been implicated in glucose/galactose malabsorption, congenital hypothyroidism, Bartter's syndrome, epilepsy, depression, autism and obsessive-compulsive disorder. Sodium-coupled transporters are blocked by a number of therapeutically important compounds, including diuretics, anticonvulsants and antidepressants, many of which have also become indispensable tools in biochemical experiments designed to probe antagonist binding sites and to elucidate transport mechanisms. Steady-state kinetic data have revealed that both competitive and noncompetitive modes of inhibition exist. Antagonist dissociation experiments on the serotonin transporter (SERT) have also unveiled the existence of a low-affinity allosteric site that slows the dissociation of inhibitors from a separate high-affinity site. Despite these strides, atomic-level insights into inhibitor action have remained elusive. Here we screen a panel of molecules for their ability to inhibit LeuT, a prokaryotic homologue of mammalian neurotransmitter sodium symporters, and show that the tricyclic antidepressant (TCA) clomipramine noncompetitively inhibits substrate uptake. Cocrystal structures show that clomipramine, along with two other TCAs, binds in an extracellular-facing vestibule about 11 Å above the substrate and two sodium ions, apparently stabilizing the extracellular gate in a closed conformation. Off-rate assays establish that clomipramine reduces the rate at which leucine dissociates from LeuT and reinforce our contention that this TCA inhibits LeuT by slowing substrate release. Our results represent a molecular view into noncompetitive inhibition of a sodium-coupled transporter and define principles for the rational design of new inhibitors.

Original languageEnglish (US)
Pages (from-to)952-956
Number of pages5
JournalNature
Volume448
Issue number7156
DOIs
StatePublished - Aug 23 2007

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Neurotransmitter Transport Proteins
Antidepressive Agents
Clomipramine
Sodium
Binding Sites
Tricyclic Antidepressive Agents
Plasma Membrane Neurotransmitter Transport Proteins
Dissociative Disorders
Bartter Syndrome
Ions
Allosteric Site
Congenital Hypothyroidism
Serotonin Plasma Membrane Transport Proteins
Aptitude
Obsessive-Compulsive Disorder
Lipid Bilayers
Autistic Disorder
Diuretics
Leucine
Anticonvulsants

ASJC Scopus subject areas

  • General

Cite this

Antidepressant binding site in a bacterial homologue of neurotransmitter transporters. / Singh, Satinder K.; Yamashita, Atsuko; Gouaux, Eric.

In: Nature, Vol. 448, No. 7156, 23.08.2007, p. 952-956.

Research output: Contribution to journalArticle

Singh, Satinder K. ; Yamashita, Atsuko ; Gouaux, Eric. / Antidepressant binding site in a bacterial homologue of neurotransmitter transporters. In: Nature. 2007 ; Vol. 448, No. 7156. pp. 952-956.
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