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Title: LeuT-Desipramine Structure Reveals How Antidepressants Block Neurotransmitter Reuptake

Abstract

Tricyclic antidepressants exert their pharmacological effect -- inhibiting the reuptake of serotonin, norepinephrine, and dopamine -- by directly blocking neurotransmitter transporters (SERT, NET, and DAT, respectively) in the presynaptic membrane. The drug-binding site and the mechanism of this inhibition are poorly understood. We determined the crystal structure at 2.9 angstroms of the bacterial leucine transporter (LeuT), a homolog of SERT, NET, and DAT, in complex with leucine and the antidepressant desipramine. Desipramine binds at the inner end of the extracellular cavity of the transporter and is held in place by a hairpin loop and by a salt bridge. This binding site is separated from the leucine-binding site by the extracellular gate of the transporter. By directly locking the gate, desipramine prevents conformational changes and blocks substrate transport. Mutagenesis experiments on human SERT and DAT indicate that both the desipramine-binding site and its inhibition mechanism are probably conserved in the human neurotransmitter transporters.

Authors:
; ; ; ; ; ;
Publication Date:
Research Org.:
Brookhaven National Laboratory (BNL) National Synchrotron Light Source
Sponsoring Org.:
Doe - Office Of Science
OSTI Identifier:
930532
Report Number(s):
BNL-80564-2008-JA
Journal ID: ISSN 0193-4511; SCEHDK; TRN: US200904%%772
DOE Contract Number:
DE-AC02-98CH10886
Resource Type:
Journal Article
Resource Relation:
Journal Name: Science; Journal Volume: 317
Country of Publication:
United States
Language:
English
Subject:
36 MATERIALS SCIENCE; ANTIDEPRESSANTS; CONFORMATIONAL CHANGES; CRYSTAL STRUCTURE; DOPAMINE; LEUCINE; MUTAGENESIS; NORADRENALINE; SEROTONIN; SUBSTRATES; national synchrotron light source

Citation Formats

Zhou,Z., Zhen, J., Karpowich, N., Goetz, R., Law, C., Reith, M., and Wang, D. LeuT-Desipramine Structure Reveals How Antidepressants Block Neurotransmitter Reuptake. United States: N. p., 2007. Web. doi:10.1126/science.1147614.
Zhou,Z., Zhen, J., Karpowich, N., Goetz, R., Law, C., Reith, M., & Wang, D. LeuT-Desipramine Structure Reveals How Antidepressants Block Neurotransmitter Reuptake. United States. doi:10.1126/science.1147614.
Zhou,Z., Zhen, J., Karpowich, N., Goetz, R., Law, C., Reith, M., and Wang, D. Mon . "LeuT-Desipramine Structure Reveals How Antidepressants Block Neurotransmitter Reuptake". United States. doi:10.1126/science.1147614.
@article{osti_930532,
title = {LeuT-Desipramine Structure Reveals How Antidepressants Block Neurotransmitter Reuptake},
author = {Zhou,Z. and Zhen, J. and Karpowich, N. and Goetz, R. and Law, C. and Reith, M. and Wang, D.},
abstractNote = {Tricyclic antidepressants exert their pharmacological effect -- inhibiting the reuptake of serotonin, norepinephrine, and dopamine -- by directly blocking neurotransmitter transporters (SERT, NET, and DAT, respectively) in the presynaptic membrane. The drug-binding site and the mechanism of this inhibition are poorly understood. We determined the crystal structure at 2.9 angstroms of the bacterial leucine transporter (LeuT), a homolog of SERT, NET, and DAT, in complex with leucine and the antidepressant desipramine. Desipramine binds at the inner end of the extracellular cavity of the transporter and is held in place by a hairpin loop and by a salt bridge. This binding site is separated from the leucine-binding site by the extracellular gate of the transporter. By directly locking the gate, desipramine prevents conformational changes and blocks substrate transport. Mutagenesis experiments on human SERT and DAT indicate that both the desipramine-binding site and its inhibition mechanism are probably conserved in the human neurotransmitter transporters.},
doi = {10.1126/science.1147614},
journal = {Science},
number = ,
volume = 317,
place = {United States},
year = {Mon Jan 01 00:00:00 EST 2007},
month = {Mon Jan 01 00:00:00 EST 2007}
}
  • LeuT serves as the model protein for understanding the relationships between structure, mechanism and pharmacology in neurotransmitter sodium symporters (NSSs). At the present time, however, there is a vigorous debate over whether there is a single high-affinity substrate site (S1) located at the original, crystallographically determined substrate site or whether there are two high-affinity substrates sites, one at the primary or S1 site and the other at a second site (S2) located at the base of the extracellular vestibule. In an effort to address the controversy over the number of high-affinity substrate sites in LeuT, one group studied the F253Amore » mutant of LeuT and asserted that in this mutant substrate binds exclusively to the S2 site and that 1 mM clomipramine entirely ablates substrate binding to the S2 site. Here we study the binding of substrate to the F253A mutant of LeuT using ligand binding and X-ray crystallographic methods. Both experimental methods unambiguously show that substrate binds to the S1 site of the F253A mutant and that binding is retained in the presence of 1 mM clomipramine. These studies, in combination with previous work, are consistent with a mechanism ofr LeuT that involves a single high-affinity substrate binding site.« less
  • No abstract prepared.
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  • Spectrin and ankyrin participate in membrane organization, stability, signal transduction, and protein targeting; their interaction is critical for erythrocyte stability. Repeats 14 and 15 of {beta}I-spectrin are crucial for ankyrin recognition, yet the way spectrin binds ankyrin while preserving its repeat structure is unknown. We have solved the crystal structure of the {beta}I-spectrin 14,15 di-repeat unit to 2.1 {angstrom} resolution and found 14 residues critical for ankyrin binding that map to the end of the helix C of repeat 14, the linker region, and the B-C loop of repeat 15. The tilt (64{sup o}) across the 14,15 linker is greatermore » than in any published di-repeat structure, suggesting that the relative positioning of the two repeats is important for ankyrin binding. We propose that a lack of structural constraints on linker and inter-helix loops allows proteins containing spectrin-like di-repeats to evolve diverse but specific ligand-recognition sites without compromising the structure of the repeat unit. The linker regions between repeats are thus critical determinants of both spectrin's flexibility and polyfunctionality. The putative coupling of flexibility and ligand binding suggests a mechanism by which spectrin might participate in mechanosensory regulation.« less