SITE OF FASCICULIN INTERACTION WITH ACETYLCHOLINESTERASE

Fasciculin, a 6750-Da peptide from the venom of Dendroaspis, is known to inhibit reversibly mammalian and fish acetylcholinesterases at pico molar concentrations, but is a relatively weak inhibitor of avian, rep tile, and insect acetylcholinesterases and mammalian butyrylcholineste rases. An examination of fasciculin association with several mutant fo rms of recombinant DNA-derived acetylcholinesterase from mouse shows t hat it interacts with a cluster of residues near the rim of the gorge on the enzyme. The aromatic residues, Trp286, Tyr72, and Tyr124, have the most marked influence on fasciculin binding, whereas Asp74, a char ged residue in the vicinity of the binding site that affects the bindi ng of low molecular weight inhibitors, has little influence on fascicu lin binding. The 3 aromatic residues are unique to the susceptible ace tylcholinesterases and, along with Asp74, constitute part of the perip heral anionic site. Fasciculin falls in the family of three-loop toxin s that include the receptor blocking alpha-toxins and cardiotoxins. Fr om this basic structural motif, a binding site has evolved on fascicul in to be highly specific for the peripheral site on acetylcholinestera se. Acetylthiocholine affects rates of fasciculin binding at concentra tions causing substrate inhibition. In the case of the mutant cholines terases where rates of fasciculin dissociation are more rapid, steady state kinetic parameters also show acetylthiocholine-fasciculin compet ition to be consistent with occupation at a peripheral or substrate in hibition site rather than the active center.