Characterisation of the heterotrimeric presynaptic phospholipase A 2 neurotoxin complex from the venom of the common death adder (Acanthophis antarcticus)

• Main Authors: Blacklow, B, Escoubas, P, Nicholson, GM
• Format: Article in Journal/Newspaper
• Language: unknown
• Published: 2010
• Subjects: Pharmacology & Pharmacy; Presynaptic Terminals; Neuromuscular Junction; Animals; Chickens; Elapidae; Receptors; Presynaptic; Elapid Venoms; Neurotoxins; Biological Assay; Chromatography; High Pressure Liquid; Spectrometry; Mass; Electrospray Ionization; Matrix-Assisted Laser Desorption-Ionization; Muscle Contraction; Phospholipases A2; Antarc*; antarcticus
• Online Access: http://hdl.handle.net/10453/13734

While Australo-Papuan death adder neurotoxicity is generally considered to be due to the actions of reversible competitive postsynaptic α-neurotoxins, the neurotoxic effects are often poorly reversed by antivenom or anticholinesterases. This suggests that the venom may contain a snake presynaptic phospholipase A2 (PLA2) neurotoxin (SPAN) that binds irreversibly to motor nerve terminals to inhibit neurotransmitter release. Using size-exclusion liquid chromatography under non-reducing conditions, we report the isolation and characterisation of a high molecular mass SPAN complex, P-elapitoxin-Aa1a (P-EPTX-Aa1a), from the venom of the common death adder Acanthophis antarcticus. Using the chick biventer-cervicis nerve-muscle preparation, P-EPTX-Aa1a (44,698Da) caused inhibition of nerve-evoked twitch contractions while responses to cholinergic agonists and KCl remained unaffected. P-EPTX-Aa1a also produced significant fade in tetanic contractions and a triphasic timecourse of neuromuscular blockade. These actions are consistent with other SPANs that inhibit acetylcholine release. P-EPTX-Aa1a was found to be a heterotrimeric complex composed of α, β and γ-subunits in a 1:1:1 stoichiometry with each subunit showing significant N-terminal sequence homology to the subunits of taipoxin, a SPAN from Oxyuranus s. scutellatus. Like taipoxin, only the α-chain produced any signs of neurotoxicity or displayed significant PLA2 enzymatic activity. Preincubation with monovalent death adder antivenom or suramin, or inhibition of PLA2 activity by incubation with 4-bromophenacyl bromide, either prevented or significantly delayed the onset of toxicity by P-EPTX-Aa1a. However, antivenom failed to reverse neurotoxicity. Early intervention with antivenom may therefore be important in severe cases of envenomation by A. antarcticus, given the presence of potent irreversible presynaptic neurotoxins. © 2010 Elsevier Inc.