Dynamics and orientation of N+(CD3)3-bromoacetylcholine bound to its binding site on the nicotinic acetylcholine receptor
- *Biomembrane Structure Unit, Biochemistry Department, University of Oxford, South Parks Road, Oxford, OX1 3QU United Kingdom; and ‡Department of Anesthesia, Massachusetts General Hospital and Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02114
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Edited by Arthur Karlin, Columbia University College of Physicians and Surgeons, New York, NY, and approved December 1, 2000 (received for review August 1, 2000)
Abstract
Dynamic and structural information has been obtained for an analogue of acetylcholine while bound to the agonist binding site on the nicotinic acetylcholine receptor (nAcChoR), using wide-line deuterium solid-state NMR. Analysis of the deuterium lineshape obtained at various temperatures from unoriented nAcChoR membranes labeled with deuterated bromoacetylcholine (BAC) showed that the quaternary ammonium group of the ligand is well constrained within the agonist binding site when compared with the dynamics observed in the crystalline solids. This motional restriction would suggest that a high degree of complementarity exists between the quaternary ammonium group of the ligand and the protein within the agonist binding site. nAcChoR membranes were uniaxially oriented by isopotential centrifugation as determined by phosphorous NMR of the membrane phospholipids. Analysis of the deuterium NMR lineshape of these oriented membranes enriched with the nAcChoR labeled with N+(CD3)3-BAC has enabled us to determine that the angle formed between the quaternary ammonium group of the BAC and the membrane normal is 42° in the desensitized form of the receptor. This measurement allows us to orient in part the bound ligand within the proposed receptor binding site.
Footnotes
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↵ † Present address: Laboratory for Physical Chemistry, Eidgenössische Technische Hochschule, Universitatestrasse 22, CH-8092 Zürich, Switzerland.
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↵ § To whom reprint requests should be addressed. E-mail: awatts{at}bioch.ox.ac.uk.
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This paper was submitted directly (Track II) to the PNAS office.
- Abbreviations:
- nAcChoR,
- nicotinic acetylcholine receptor;
- ACh,
- acetylcholine;
- BAC,
- bromoacetylcholine
- Copyright © 2001, The National Academy of Sciences
