Title: Crystal Structures of a Cysteine-modified Mutant in Loop D of Acetylcholine-binding Protein
Authors: Brams, Marijke
Gay, Elaine A
Saez, Jose Colon
Guskov, Albert
van Elk, Rene
van der Schors, Roel C
Peigneur, Steve
Tytgat, Jan
Strelkov, Sergei
Smit, August B
Yakel, Jerrel L
Ulens, Chris # ×
Issue Date: Feb-2011
Publisher: American Society for Biochemistry and Molecular Biology
Series Title: Journal of Biological Chemistry vol:286 issue:6 pages:4420-4428
Abstract: Covalent modification of alpha 7 W55C nicotinic acetylcholine receptors (nAChR) with the cysteine-modifying reagent [2-(trimethylammonium) ethyl] methanethiosulfonate (MTSET+) produces receptors that are unresponsive to acetylcholine, whereas methyl methanethiolsulfonate (MMTS) produces enhanced acetylcholine-gated currents. Here, we investigate structural changes that underlie the opposite effects of MTSET+ and MMTS using acetylcholine-binding protein (AChBP), a homolog of the extracellular domain of the nAChR. Crystal structures of Y53C AChBP show that MT-SET+-modification stabilizes loop C in an extended conformation that resembles the antagonist-bound state, which parallels our observation that MTSET+ produces unresponsive W55C nAChRs. The MMTS-modified mutant in complex with acetylcholine is characterized by a contracted C-loop, similar to other agonist-bound complexes. Surprisingly, we find two acetylcholine molecules bound in the ligand-binding site, which might explain the potentiating effect of MMTS modification in W55C nAChRs. Unexpectedly, we observed in the MMTS-Y53C structure that ten phosphate ions arranged in two rings at adjacent sites are bound in the vestibule of AChBP. We mutated homologous residues in the vestibule of alpha 1 GlyR and observed a reduction in the single channel conductance, suggesting a role of this site in ion permeation. Taken together, our results demonstrate that targeted modification of a conserved aromatic residue in loop D is sufficient for a conformational switch of AChBP and that a defined region in the vestibule of the extracellular domain contributes to ion conduction in an-ion-selective Cys-loop receptors.
ISSN: 0021-9258
Publication status: published
KU Leuven publication type: IT
Appears in Collections:Biocrystallography
Laboratory of Structural Neurobiology
Toxicology and Pharmacology
× corresponding author
# (joint) last author

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