Characterisation of the heterotrimeric presynaptic phospholipase A2 neurotoxin complex from the venom of the common death adder (Acanthophis antarcticus) |
| |
Authors: | Benjamin Blacklow Graham M. Nicholson |
| |
Affiliation: | a Neurotoxin Research Group, Department of Medical & Molecular Biosciences, University of Technology, Sydney, P.O. Box 123, Broadway, NSW 2007, Australia b Université de Nice - Sophia Antipolis, Institut de Pharmacologie Moléculaire et Cellulaire (IPMC), CNRS UMR6097, 660 Route des Lucioles, 06560 Valbonne, France |
| |
Abstract: | 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 (44,698 Da) 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. |
| |
Keywords: | ACh, acetylcholine ANOVA, analysis of variance 4BPB, 4-bromophenacyl bromide CBCNM, chick biventer-cervicis nerve-muscle CCh, carbachol ESI-Q-TOF, electrospray ionisation quadrupole time-of-flight MALDI-TOF, matrix-assisted laser desorption ionisation time-of-flight NSW, New South Wales PLA2, phospholipase A2 P-EPTX-Aa1a, P-elapitoxin-Aa1a RP-HPLC, reverse-phase high-pressure liquid chromatography SPAN, snake presynaptic phospholipase A2 neurotoxin TFA, trifluoroacetic acid t90, time to 90% neuromuscular blockade Ve, elution volume Vo, void volume |
本文献已被 ScienceDirect 等数据库收录! |
|