Interaction of the neurotoxic and nontoxic secretory phospholipases A2 with the crotoxin inhibitor from Crotalus serum

Crotalus durissus terrificus snakes possess a protein in their blood, named crotoxin inhibitor from Crotalus serum (CICS), which protects them against crotoxin, the main toxin of their venom. CICS neutralizes the lethal potency of crotoxin and inhibits its phospholipase A(2) (PLA(2)) activity. The aim of the present study is to investigate the specificity of CICS towards snake venom neurotoxic PLA(2)s (beta-neurotoxins) and nontoxic mammalian PLA(2)s. This investigation shows that CICS does not affect the enzymatic activity of pancreatic and nonpancreatic PLA(2)s, bee venom PLA(2) and Elapidae beta-neurotoxins but strongly inhibits the PLA(2) activity of Viperidae beta-neurotoxins. Surface plasmon resonance and PAGE studies further demonstrated that CICS makes complexes with monomeric and multimeric Viperidae beta-neurotoxins but does not interact with nontoxic PLA(2)s. In the case of dimeric beta-neurotoxins from Viperidae venoms (crotoxin, Mojave toxin and CbICbII), which are made by the noncovalent association of a PLA(2) with a nonenzymatic subunit, CICS does not react with the noncatalytic subunit, instead it binds tightly to the PLA(2) subunit and induces the dissociation of the heterocomplex. In vitro assays performed with Torpedo synaptosomes showed a protective action of CICS against Viperidae beta-neurotoxins but not against other PLA(2) neurotoxins, on primary and evoked liberation of acetylcholine. In conclusion, CICS is a specific PLA(2) inhibitor of the beta-neurotoxins from the Viperidae family.