Journal of Neuroscience, Vol 7, 2547-2555, Copyright © 1987 by Society for Neuroscience
Affinity labeling of neuronal acetylcholine receptor subunits with an alpha-neurotoxin that blocks receptor function
SW Halvorsen and DK Berg
An alpha-neurotoxin, Bgt 3.1, has previously been shown to recognize the
functional acetylcholine receptor (AChR) on chick autonomic neurons, since
it specifically blocks receptor function and it binds to a class of sites
on the neurons with the pharmacology, kinetics, and affinity expected for
the receptor. A monoclonal antibody, mAb 35, to the main immunogenic region
of muscle and electric organ AChR alpha subunit cross-reacts with a
component on chick autonomic neurons that, from several lines of evidence,
also appears to be the functional AChR. The identity of the
antibody-binding component has remained in doubt, however, because previous
studies indicated that in at least one instance a substantial discrepancy
existed between the number of functional AChRs estimated physiologically
and the number of putative AChRs detected by mAb 35 on the neurons. The
present findings demonstrate that Bgt 3.1 and mAb 35 recognize the same
AChRs on the neurons, and provide information about the stoichiometry of
binding and the identity of subunits associated with active sites on the
receptor. Chick ciliary ganglion neurons examined under a variety of growth
and regulatory conditions in culture displayed a constant ratio of about
2:1 for mAb 35 and Bgt 3.1 binding to cell surface sites. Treatment of the
cells with mAb 35 induced a substantial decrease in the number of Bgt 3.1
sites and vice versa. AChRs that had been covalently labeled with a
photoaffinity derivative of 125I-Bgt 3.1 in situ and then solubilized were
specifically immune-precipitated by mAb 35, demonstrating unequivocally
that the receptors possessed both Bgt 3.1 and mAb 35 binding
sites.(ABSTRACT TRUNCATED AT 250 WORDS)
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