Postsynaptic Effects of Crotoxin and of Its Isolated Subunits

Abstract

Crotoxin is a potent neurotoxin from the venom of Crotalus durissus terrificus. It is composed of two subunits: a basic phospholipase A2 with low toxicity (component B) and an acidic protein seemingly devoid of intrinsic biological activity (component A). Crotoxin and its isolated phospholipase subunit block the depolarisation caused by cholinergic agonists on the isolated electroplaque from Electrophorus electricus. The other component, which is inactive when applied alone, enhances the pharmacological activity of the phospholipase when the two components are used together. Crotoxin also blocks the increase of z2Na+ efflux caused by carbamylcholine from excitable microsacs prepared from Torpedo marmorata electric organ. Crotoxin therefore acts postsynaptically, but does not interfere with the binding of a-toxin from Nu@ nigricollis to the nicotinic cholinergic receptor site. Instead, like local anesthetics, it stabilizes a desensitized form of the acetylcholine receptor characterized by its high affinity for agonists. The phospholipase component B binds in a nonsaturable manner to receptor-rich membranes. In contrast, component A does not bind to acetylcholine receptor-rich membranes, but completely prevents the non-saturable binding of component B. When the two components are applied together, a saturable binding of the latter is observed with the acetylcholine receptor-rich membranes.Crotoxin, the major component of the brazilian rattle-snake (Crotalus durissus terrificus) venom is a very potent neurotoxin which possesses a phospholipase activity [l]. It is a complex of two distinct protein subunits [2,3] : a strongly basic component (component B, M , 12000) which carries the phospholipase activity and an acidic protein (component A, M , 10000) which has no enzymatic activity and is not toxic by itself. Whereas the isolated subunits are either not or only a little toxic, they recombine into a complex which exhibits the same toxicity as the original material [4,5].The lethal effect of crotoxin has generally been attributed to a presynaptic blockage of the neuromuscular transmission [6]. At this level, it causes a reduction of acetylcholine release by the nerve terminals similar to that observed with /l-bungarotoxin [7], notexin [8,9] and taipoxin [lo]. However, having observed a depression of the response to acetylcholine of the isolated denervated rat diaphragm, VitalBrazil [ l l ] concluded that crotoxin also acts at the postsynaptic level in the mammals. In order to examine a possible postsynaptic action of crotoxin without interference with its presynaptic action, we studied the effect of crotoxin on two postsynapticEnzyme. Phospholipase A2 (EC 3.1.1.4) preparations : the isolated electroplaque from Electrophorus electricus and the acetylcholine receptor-rich membranes from Torpedo marmorata. Crotoxin affects the response of both preparations to cholinergic agonists, and our observations suggest how interactions between the two subunits increase the pharmacological effect of the phospholipase component...

Reconstitution of a Functional Acetylcholine Receptor

“…A direct interaction of sodium cholate with the local anesthetic binding site thus cannot be excluded, but an indirect, or 'allosteric' effect of sodium cholate appears more plausible. Accordingly, it would resemble that observed with other effectors such as several phospholipases [25,46,49,51,52], general anesthetics or alcohols [44] which also stabilize the receptor in a high-affinity state and are known to perturbate the lipid phase.…”

A Molecular Mechanism for Lys49-Phospholipase A2 Activity Based on Ligand-induced Conformational Change

“…The lysophospholipids on the other hand are involved in cell signaling and phospholipid remodeling and are associated with membrane perturbation (9,10). Some members of the PLA 2 superfamily also present a wide variety of pharmacological effects, such as pre-and post-synaptic neurotoxicity (11,12), myotoxicity (13)(14)(15)(16)(17)(18)(19), cardiotoxicity (20), and platelet aggregation (21,22) among others.…”

The origin of the diversity of crotoxin isoforms in the venom of Crotalus durissus terrificus

“…Crotoxin is composed of two non-covalently associated subunits [2,31 ; a basic and weakly toxic phospholipase A,, subunit B (CB) [4,51 and an acidic protein, subunit A (CA) [6, 71, devoid of enzymic activity and non toxic. CA enhances the capacity of CB to reach its target at neuromuscular junctions and, therefore, its toxicity [8]. Crotoxin is, in fact, a mixture of variants [9-131 deriving from the combination of subunit isoforms .…”