TY - JOUR
T1 - Residues at the subunit interfaces of the nicotinic acetylcholine receptor that contribute to α-conotoxin M1 binding
AU - Sugiyama, Naoya
AU - Marchot, Pascale
AU - Kawanishi, Chiaki
AU - Osaka, Hitoshi
AU - Molles, Brian
AU - Sine, Steven M.
AU - Taylor, Palmer
PY - 1998/4
Y1 - 1998/4
N2 - The two binding sites in the pentameric nicotinic acetylcholine receptor of subunit composition α2βγδ are formed by nonequivalent α-γ and α- δ subunit interfaces, which produce site selectivity in the binding of agonists and antagonists. We show by sedimentation analysis that 125I-α- conotoxin M1 binds with high affinity to the α-δ subunit dimers, but not to α-γ dimers, nor to α, γ, and δ monomers, a finding consistent with α- conotoxin M1 selectivity for the αδ interface in the intact receptor measured by competition against α-bungarotoxin binding. We also extend previous identification of α-conotoxin M1 determinants in the γ and δ subunits to the α subunit interface by mutagenesis of conserved residues in the α subunit. Most mutations of the α subunit affect affinity similarly at the two sites, but Tyr93Phe, Val188Lys, Tyr190Thr, Tyr198Thr, and Asp152Asn affect affinity in a site-selective manner. Mutant cycle analysis reveals only weak or no interactions between mutant a and non-α subunits, indicating that side chains of the α subunit do not interact with those of the γ or δ subunits in stabilizing α-conotoxin M1. The overall findings suggest different binding configurations of α-conotoxin M1 at the α-δ and α-γ binding interfaces.
AB - The two binding sites in the pentameric nicotinic acetylcholine receptor of subunit composition α2βγδ are formed by nonequivalent α-γ and α- δ subunit interfaces, which produce site selectivity in the binding of agonists and antagonists. We show by sedimentation analysis that 125I-α- conotoxin M1 binds with high affinity to the α-δ subunit dimers, but not to α-γ dimers, nor to α, γ, and δ monomers, a finding consistent with α- conotoxin M1 selectivity for the αδ interface in the intact receptor measured by competition against α-bungarotoxin binding. We also extend previous identification of α-conotoxin M1 determinants in the γ and δ subunits to the α subunit interface by mutagenesis of conserved residues in the α subunit. Most mutations of the α subunit affect affinity similarly at the two sites, but Tyr93Phe, Val188Lys, Tyr190Thr, Tyr198Thr, and Asp152Asn affect affinity in a site-selective manner. Mutant cycle analysis reveals only weak or no interactions between mutant a and non-α subunits, indicating that side chains of the α subunit do not interact with those of the γ or δ subunits in stabilizing α-conotoxin M1. The overall findings suggest different binding configurations of α-conotoxin M1 at the α-δ and α-γ binding interfaces.
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U2 - 10.1124/mol.53.4.787
DO - 10.1124/mol.53.4.787
M3 - Article
C2 - 9547372
AN - SCOPUS:0031967095
SN - 0026-895X
VL - 53
SP - 787
EP - 794
JO - Molecular pharmacology
JF - Molecular pharmacology
IS - 4
ER -