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Mapping Residues in the Ligand-Binding Domain of the 5-HT3 Receptor onto d-Tubocurarine Structure
Journal article   Peer reviewed

Mapping Residues in the Ligand-Binding Domain of the 5-HT3 Receptor onto d-Tubocurarine Structure

Dong Yan, Julia K. Meyer and Michael M. White
Molecular pharmacology, v 70(2), pp 571-578
01 Aug 2006
PMID: 16723497

Abstract

The serotonin 5-HT 3 receptor (5-HT 3 R) is a member of the cys-loop ligand-gated ion channel family. We have used the combination of site-directed mutagenesis, homology modeling of the 5-HT 3 R extracellular domain, and ligand docking simulations as a way to map the architecture of the 5-HT 3 R ligand binding domain. Mutation of Phe226 in loop C of the binding site to tyrosine (F226Y) has no effect on the apparent affinity of the competitive antagonist d -tubocurarine ( d TC) for the receptor. On the other hand, replacement of Asn128 in loop A of the binding site with alanine (N128A) increases the apparent affinity of d TC by approximately 10-fold. Double-mutant cycle analysis employing a panel of d TC analogs with substitutions at various positions to identify specific points of interactions between the d TC analogs and Asn128 suggests that Asn128 makes a direct interaction with the 2′N of d TC. Molecular modeling of the 5-HT 3 R extracellular domain using the antagonist-bound conformation of the Aplysia californica acetylcholine binding protein as a template followed by ligand docking simulations produces two classes of structures of the 5-HT 3 R/ d TC complex; only one of these has the 2′N of d TC positioned at Asn128 and thus is consistent with the data from this study and previously published data. The use of the rigid d TC analogs as “molecular rulers” in conjunction with double-mutant cycle analysis of mutant receptors can allow the spatial mapping of the position of various residues in the ligand-binding site.

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