PRESYNAPTICALLY ACTING SNAKE-VENOM PHOSPHOLIPASE A(2) ENZYMES ATTACK UNIQUE SUBSTRATES

Synaptosomes were incubated with bovine serum albumin (BSA) to examine whether the presynaptic action of snake venom phospholipase A(2) (PLA (2)) toxins is due either to the release of fatty acids resistant to e xtraction by BSA or to the liberation of a specific fatty acid type. I n the presence of BSA (0.5% or 1.0%) two PLA(2) enzymes from Naja naja atra and Naja naja kaouthia snake venoms that do not have a predomina nt presynaptic action at the neuromuscular junction (PS-) did not stim ulate acetylcholine (ACh) release from synaptosomes. In contrast, two PLA(2) enzymes (beta-bungarotoxin, scutoxin) that do have a predominan t presynaptic action at the neuromuscular junction (PS+) did stimulate ACh release. BSA did not antagonize PS- enzymes by more efficiently e xtracting the fatty acids produced by these enzymes relative to PS+ en zymes. While absolute amounts of total and unsaturated fatty acid prod uced overlapped for the PS- and PS+ enzymes, the two PS+ enzymes produ ced a significantly greater absolute amount and relative percentage of palmitic acid (16:0) than did either of the PS- enzymes. However, the levels of free palmitic acid remaining in the synaptosomes where they would exert effects on ACh release were similar for the N. n. kaouthi a PLA(2) (PS-) and beta-bungarotoxin (PS+). Therefore, the total (supe rnatant plus synaptosomal) amount of palmitic acid produced per se did not account for stimulation of ACh release, since the greater amounts produced by the PS+ enzymes were removed from the synaptosomes by BSA . The production of higher levels of palmitic acid suggests either tha t PS+ enzymes gain access to sites containing phospholipid substrates unavailable to the PS- enzymes, or that they have a different substrat e preference. These findings suggest new possibilities for the mechani sm of PS+PLA(2) action, including site-directed enzymatic activity and protein acylation.