Open AccessPhotoaffinity labeling of Torpedo acetylcholine receptor by physostigmine
Author(s) -
SCHRATTENHOLZ Andre,
GODOVACZIMMERMANN Jasminka,
SCHÄFER HansJochen,
ALBUQUERQUE Edson X.,
MAELICKE Alfred
Publication year - 1993
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1993.tb18187.x
Subject(s) - physostigmine , torpedo , acetylcholine , acetylcholine receptor , chemistry , nicotinic acetylcholine receptor , binding site , muscarinic acetylcholine receptor , cholinesterase , nicotinic agonist , biochemistry , muscarinic acetylcholine receptor m5 , receptor , biophysics , muscarinic acetylcholine receptor m3 , pharmacology , biology
The plant alkaloid physostigmine, an established anti‐cholinesterase agent of the carbamate type, has recently been shown to bind to the nicotinic acetylcholine receptor from Torpedo marmorata electrocytes [Okonjo, K. O., Kuhlmann, J. & Maelicke, A. (1991) Eur. J. Biochem. 200 , 671–677]. Pharmacological studies of physostigmine‐induced ion flux into nicotinic‐acetylcholine‐receptor‐rich membrane vesicles, indicated distinct binding sites for physostigmine and acetylcholine. As shown in this study by photoaffinity labeling with [phenyl‐(n)‐ 3 H](‐)physostigmine, the physostigmine‐binding site is located within the same subunit (α polypeptide) of the receptor as the acetylcholine‐binding site. Using a variety of proteolytic cleavage conditions for the purified α polypeptide, several [ 3 H]physostigmine‐labeled peptides were isolated and sequenced. From the radioactivity released in the course of the Edman degradations of the labeled peptides, it was found that the label was associated in all cases with Lys125. These results identify a novel ligand‐binding site for the Torpedo nicotinic acetylcholine receptor that is different in location from binding sites identified previously for acetylcholine, its established agonists and antagonists, and direct channel blockers.