SLC6 Neurotransmitter Transporters: Structure, Function, and Regulation

Kristensen, Anders S.; Andersen, Jacob; Jørgensen, Trine N.; Sørensen, Lena; Eriksen, Jacob; Løland, Claus J.; Strømgaard, Kristian; Gether, Ulrik

doi:10.1124/pr.108.000869

Cited by 707 publications

(802 citation statements)

References 705 publications

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“…They are also the primary drug target of clinical antidepressants such as Prozac, tricyclic antidepressants, sertraline and fluoxetine [3,4] and illegal drugs that include cocaine and amphetamines [5]. As the only NSS homologue amenable to high-resolution structural studies, LeuT [6] is a paradigm for both sodium-dependent and -independent transporters [7,8], and is most importantly a molecular blueprint for structural and functional studies of eukaryotic NSSs [9][10][11][12].…”

Section: Introductionmentioning

confidence: 99%

Substrate binds in the S1 site of the F253A mutant of LeuT, a neurotransmitter sodium symporter homologue

Wang

Gouaux

2012

EMBO Reports

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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 F253A 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 for LeuT that involves a single high-affinity substrate binding site.

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Section: Introductionmentioning

confidence: 99%

Substrate binds in the S1 site of the F253A mutant of LeuT, a neurotransmitter sodium symporter homologue

Wang

Gouaux

2012

EMBO Reports

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“…The role of uptake1 or 'reuptake' transporters in the clearance of dopamine, serotonin, and norepinephrine at the synaptic cleft has been well established for many years (Daws, 2009;Kristensen et al, 2011). These biogenic amine neurotransmitter transporters are classified according to their affinity/capacity for substrates/ inhibitors and sensitivity to ions.…”

Section: Introductionmentioning

confidence: 99%

Transport of Biogenic Amine Neurotransmitters at the Mouse Blood–Retina and Blood–Brain Barriers by Uptake1 and Uptake2

André

Saubaméa

Cochois-Guégan

et al. 2012

J Cereb Blood Flow Metab

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Uptake1 and uptake2 transporters are involved in the extracellular clearance of biogenic amine neurotransmitters at synaptic clefts. We looked for them at the blood-brain barrier (BBB) and bloodretina barriers (BRB), where they could be involved in regulating the neurotransmitter concentration and modulate/terminate receptor-mediated effects within the neurovascular unit (NVU). Uptake2 (Oct1-3/ Slc22a1-3, Pmat/Slc29a4) and Mate1/Slc47a1 transporters are also involved in the transport of xenobiotics. We used in situ carotid perfusion of prototypic substrates like-serotonin, and [ 3 H]-dopamine, changes in ionic composition and genetic deletion of Oct1-3 carriers to detect uptake1 and uptake2 at the BBB and BRB. We showed that uptake1 and uptake2 are involved in the transport of [ 3 H]-dopamine and [ 3 H]-MPP + at the blood luminal BRB, but not at the BBB. These functional studies, together with quantitative RT-PCR and confocal imaging, suggest that the mouse BBB lacks uptake1 (Net/Slc6a2, Dat/Slc6a3, Sert/Slc6a4), uptake2, and Mate1 on both the luminal and abluminal sides. However, we found evidence for functional Net and Oct1 transporters at the luminal BRB. These heterogeneous transport properties of the brain and retina NVUs suggest that the BBB helps protect the brain against biogenic amine neurotransmitters in the plasma while the BRB has more of a metabolic/endocrine role.

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“…The S1 site is highly conserved in mammalian transporters, and mutagenesis of many DAT residues near this site, including Phe-76, Asp-79, Val-152, Phe-155, Tyr-156, Asn-157, Phe-319, Val-327, and Ser-421, reduces dopamine transport (31)(32)(33)(34)(35)(36)(37)(38), supporting the participation of these amino acids in substrate recognition or translocation. In particular, Asp-79 in TM1 of DAT and the homologous Asp residues in NET and SERT coordinate the positive charge on monoamine substrates and play an essential role in transport (6,12,13,15,39,40). However, a second, highly controversial substrate site (S2) proposed to play a role in the initiation of transport has been identified in LeuT in the extracellular vestibule above the outer gate (27,(41)(42)(43).…”

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confidence: 99%

“…DAT and the related cocaine-sensitive norepinephrine and serotonin transporters (NET and SERT) belong to the neurotransmitter sodium symporter (NSS) subfamily of solute carrier 6 transporters (SLC6A) that share a common topology of 12 transmembrane (TM) domains connected by extracellular (EL) and intracellular (IL) loops (6,12,13). Solute translocation is driven by a Na ϩ -and Cl Ϫ -dependent alternating access mechanism in which the proteins cycle through outwardly and inwardly facing conformations that bind and release substrates on opposite sides of the membrane (14).…”

mentioning

confidence: 99%

Computational and Biochemical Docking of the Irreversible Cocaine Analog RTI 82 Directly Demonstrates Ligand Positioning in the Dopamine Transporter Central Substrate-binding Site

Dahal

Pramod

Sharma

et al. 2014

Journal of Biological Chemistry

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Background: Cocaine interaction with DAT was assessed using the irreversible binding cocaine analog RTI 82. Results: Molecular modeling and peptide mapping identify adduction of RTI 82 to Phe-319 and Phe-320 of rat DAT and human DAT, respectively. Conclusion: Tropane-based pharmacophores bind to DAT in the central substrate site. Significance: Mapping the cocaine-binding site reveals new insights for medication discovery.

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SLC6 Neurotransmitter Transporters: Structure, Function, and Regulation

Cited by 707 publications

References 705 publications

Substrate binds in the S1 site of the F253A mutant of LeuT, a neurotransmitter sodium symporter homologue

Substrate binds in the S1 site of the F253A mutant of LeuT, a neurotransmitter sodium symporter homologue

Transport of Biogenic Amine Neurotransmitters at the Mouse Blood–Retina and Blood–Brain Barriers by Uptake1 and Uptake2

Computational and Biochemical Docking of the Irreversible Cocaine Analog RTI 82 Directly Demonstrates Ligand Positioning in the Dopamine Transporter Central Substrate-binding Site

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