MODEL FOR THE INTERACTION OF CROTOXIN, A PHOSPHOLIPASE-A(2) NEUROTOXIN, WITH PRESYNAPTIC MEMBRANES

Crotoxin is a phospholipase A2 neurotoxin that impairs the release of acetylcholine at neuromuscular junctions, primarily at the presynaptic level. It associates a phospholipase A2 subunit, CB, with a chaperon subunit, CA. We have shown elsewhere that the purely cholinergic synap tosomes from the Torpedo electric organ provided a convenient model to study the pharmacology of crotoxin and other related neurotoxins [Del ot, E., & Bon, C.(1992) J. Neurochem. 58, 311-319]. In the present exp eriments, we labeled crotoxin with I-125 and demonstrated saturable bi nding to Torpedo presynaptic membranes. In the range of concentrations that was effective on synaptosomes, crotoxin bound to a single class of sites without cooperativity. The binding was affected by divalent c ations, and its kinetics was rather complex. We observed a competition between crotoxin and related neurotoxins, but not CB. Although CA cou ld not bind, it could compete efficiently with crotoxin. We compare ou r results with those previously obtained by others on guinea pig brain membranes. On Torpedo membranes, as on all models tested, CB was the major species bound to the membrane, while CA remained in solution. Ho wever, the mechanism underlying this phenomenon had never been clarifi ed. The major question is the time scale of the events, i.e., does CB bind before or after dissociating from CA? Our results indicate that t he predominant pathway involves the formation of a ternary complex bet ween crotoxin's subunits and the acceptor site preceding the release o f CA.