Identification of residues at the α and ε subunit interfaces mediating species selectivity of Waglerin-1 for nicotinic acetylcholine receptors

Brian E. Molles, Parastoo Rezai, Eric F. Kline, Joseph J. McArdle, Steven M Sine, Palmer Taylor

Research output: Contribution to journalArticle

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Abstract

Waglerin-1 (Wtx-1) is a 22-amino acid peptide that is a competitive antagonist of the muscle nicotinic receptor (nAChR). We find that Wtx-1 binds 2100-fold more tightly to the α-ε than to the α-δ binding site interface of the mouse nAChR. Moreover, Wtx-1 binds 100-fold more tightly to the α-ε interface from mouse nAChR than that from rat or human sources. Site-directed mutagenesis of residues differing in the extracellular domains of rat and mouse ε subunits indicates that residues 59 and 115 mediate the species difference in Wtx-1 affinity. Mutation of residues 59 (Asp in mouse, Glu in rat ε) and 115 (Tyr in mouse, Ser in rat ε) converts Wtx-1 affinity for the α-ε interface of one species to that of the other species. Studies of different mutations at position 59 indicate both steric and electrostatic contributions to Wtx-1 affinity, whereas at position 115, both aromatic and polar groups contribute to affinity. The human nAChR also has lower affinity for Wtx-1 than mouse nAChR, but unlike rat nAChR, residues in both α and ε subunits mediate the affinity difference. In human nAChR, polar residues (Ser-187 and Thr-189) confer low affinity, whereas in mouse nAChR aromatic residues (Trp-187 and Phe-189) confer high affinity. The overall results show that non-conserved residues at the nAChR binding site, although not crucial for activation by ACh, govern the potency of neuromuscular toxins.

Original languageEnglish (US)
Pages (from-to)5433-5440
Number of pages8
JournalJournal of Biological Chemistry
Volume277
Issue number7
DOIs
StatePublished - Feb 15 2002
Externally publishedYes

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Nicotinic Receptors
Rats
Binding Sites
Mutagenesis
Mutation
Muscle
Electrostatics
Chemical activation
Site-Directed Mutagenesis
waglerin
Static Electricity
Amino Acids
Peptides
Muscles

ASJC Scopus subject areas

  • Biochemistry

Cite this

Identification of residues at the α and ε subunit interfaces mediating species selectivity of Waglerin-1 for nicotinic acetylcholine receptors. / Molles, Brian E.; Rezai, Parastoo; Kline, Eric F.; McArdle, Joseph J.; Sine, Steven M; Taylor, Palmer.

In: Journal of Biological Chemistry, Vol. 277, No. 7, 15.02.2002, p. 5433-5440.

Research output: Contribution to journalArticle

Molles, Brian E. ; Rezai, Parastoo ; Kline, Eric F. ; McArdle, Joseph J. ; Sine, Steven M ; Taylor, Palmer. / Identification of residues at the α and ε subunit interfaces mediating species selectivity of Waglerin-1 for nicotinic acetylcholine receptors. In: Journal of Biological Chemistry. 2002 ; Vol. 277, No. 7. pp. 5433-5440.
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