A second extracellular site is required for norepinephrine transport by the human norepinephrine transporter

Wang, Ching-I A., Shaikh, Nausad H., Ramu, Soumya and Lewis, Richard J. (2012) A second extracellular site is required for norepinephrine transport by the human norepinephrine transporter. Molecular Pharmacology, 82 5: 898-909. doi:10.1124/mol.112.080630


Author Wang, Ching-I A.
Shaikh, Nausad H.
Ramu, Soumya
Lewis, Richard J.
Title A second extracellular site is required for norepinephrine transport by the human norepinephrine transporter
Journal name Molecular Pharmacology   Check publisher's open access policy
ISSN 0026-895X
1521-0111
Publication date 2012-11-01
Sub-type Article (original research)
DOI 10.1124/mol.112.080630
Volume 82
Issue 5
Start page 898
End page 909
Total pages 12
Place of publication Bethesda, MD, United States
Publisher American Society for Pharmacology and Experimental Therapeutics
Collection year 2013
Language eng
Abstract The human norepinephrine transporter (NET) is implicated in many neurological disorders and is a target of tricyclic antidepressants and nisoxetine (NX). We used molecular docking simulations to guide the identification of residues likely to affect substrate transport and ligand interactions at NET. Mutations to alanine identified a hydrophobic pocket in the extracellular cavity of NET, comprising residues Thr80, Phe317, and Tyr317, which was critical for efficient norepinephrine (NE) transport. This secondary NE substrate site (NESS-2) overlapped the NX binding site, comprising Tyr84, Phe317, and Tyr317, and was positioned ∼11 Å extracellular to the primary site for NE (NESS-1). Thr80 in NESS-2 appeared to be critical in positioning NE for efficient translocation to NESS-1. Three residues identified as being involved in gating the reverse transport of NE (Arg81, Gln314, and Asp473) did not affect NE affinity for NESS-1. Mutating residues adjacent to NESS-2 abolished NET expression (D75A and L76A) or appeared to affect NET folding (S419A), suggesting important roles in stabilizing NET structure, whereas W308A and F388A at the top of NESS-2 abolished both NE transport and NX binding. Our findings are consistent with a multistep model of substrate transport by NET, for which a second, shallow extracellular NE substrate site (NESS-2) is required for efficient NE transport by NET.
Keyword Human dopamine transporter
Serotonin transporter
Neurotransmitter transporters
Binding-site
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ
Additional Notes Published online before print August 8, 2012.

Document type: Journal Article
Sub-type: Article (original research)
Collections: Official 2013 Collection
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