Labeling studies of photolabile philanthotoxins with nicotinic acetylcholine receptors: mode of interaction between toxin and receptor Original Research Article

Background: The nicotinic acetylcholine receptors (nAChRs) and glutamate receptors are ligand-gated cation channels composed of five separate polypeptide chains. A 43 kDa protein of unknown function is noncovalently associated with the cytoplasmic side of nAChR in vivo. The -venoms of many wasps and spiders ccntain toxins that block the activity of these channels. Philanthotoxin-433 (PhTX-433) is a non-competitive channel blocker found in the venom of the wasp Philanthus. We have used a photolabile derivative to investigate how PhTX-433 interacts with nAChRs. Results: A radiolabeled PhTX analog, containing a photolabile group substituted on one of its aromatic rings, photocrosslinked to all five subunits (α, α′, β, γ, δ) of purified nAChR in the absence of the 43 kDa protein. In the presence of the 43 kDa protein, the a subunit was preferentially labeled. Proteolysis of the receptor after crosslinking indicated that the hydrophobic end (head) of the PhTx-433 analog bound to the cytoplasmic loop(s) of the a-subunit. Binding is inhibited by other non-competitive channel blockers such as the related polyamine-amide toxins from spiders and chlorpromazine.