Two Novel α-Neurotoxins Isolated from the Taipan Snake, Oxyuranus scutellatus, Exhibit Reduced Affinity for Nicotinic Acetylcholine Receptors in Brain and Skeletal Muscle

Abstract

Three novel toxic peptides were purified to homogeneity from the venom of the Australian taipan snake, Oxyuranus scutellatus scutellatus. On the basis of complete amino acid sequence analyses, two of these toxins belong to the family of short-chain alpha-neurotoxins found in elapid and hydrophid snake venoms and are the first postsynaptic neurotoxins identified in taipan venom. Radioligand binding studies confirm that taipan toxins 1 and 2 inhibit the binding of [125I]-alpha-bungarotoxin to nicotinic acetylcholine receptors in skeletal muscle with IC50 values of 2.4-2.5 nM but are 5-fold less potent in this assay than alpha-bungarotoxin or the two short-chain alpha-neurotoxins erabutoxin a and erabutoxin b. Taipan toxins 1 and 2 do not antagonize [125I]-alpha-bungarotoxin binding to central neuronal nicotinic receptors at concentrations up to 3 microM. We find that erabutoxin a and erabutoxin b do inhibit the binding of [125I]-alpha-bungarotoxin to central neuronal nicotinic receptors but are over 350-fold less potent than long-chain alpha-neurotoxins at these receptors. The novel alpha-neurotoxins from taipan venom do not inhibit the binding of [3H]nicotine to high-affinity nicotine receptors in brain, a property they share with alpha-bungarotoxin and the erabutoxins. The results demonstrate that at least two neuromuscular junction-blocking peptides are present in taipan venom. Nonconservative substitutions at position 32 in both taipan toxin 1 and 2 may be responsible for the observed decreases in affinities of the toxins of 5-fold for muscle receptors (compared to alpha-bungarotoxin) and over 10-fold for alpha-bungarotoxin-sensitive nicotinic receptors in brain (compared to the structurally similar short-chain alpha-neurotoxins erabutoxin a and erabutoxin b).

Only Snake Curaremimetic Toxins with a Fifth Disulfide Bond Have High Affinity for the Neuronal α7 Nicotinic Receptor

“…This is in sharp contrast with the Torpedo receptor, which recognizes with high affinities both short chain and long chain curaremimetic toxins from snake venoms. A differential selectivity between long chain and short chain toxins has been depicted previously toward the chick optic lobe (34), suggesting that our observations, made with a recombinant ␣7 receptor, may also occur in vivo. However, for this extrapolation to be valid, one needs to demonstrate that the recombinant chimeric receptor behaves just like the natural ␣7 receptor toward the toxins.…”

“…Recently, it was observed that ␣-Bgtx and two new short chain toxins from the Australian snake Oxyuranus scutellatus bind to homogenates from chick optic lobes, a tissue that is likely to contain various neuronal receptors, including the ␣7 receptor. How- ever, only the long chain toxin, ␣-Bgtx, binds to the homogenate with high affinity (34).…”

Distinct activities of novel neurotoxins from Australian venomous snakes for nicotinic acetylcholine receptors

“…We described here the identification of a neurotoxin from the venom gland cDNA library of O. scutellatus, and its further characterisation by 5RACE and cloning. The transcript was identified as a short-chain neurotoxin which was previously described at the mature protein level from the venom of the coastal taipan [19]. Subsequent PCR amplification identified a related family of toxins within the venom glands of eight other Australian elapids, including the short neurotoxin Pa-a from P. australis, which shares 72.6% identity at the protein level (Fig.…”

“…The mature protein sequence of Os SNTX-1, excluding propeptide, had previously been determined by Zamudio et al [19], who described the isolation and characterisation of two toxins, toxins 1 and 2, from the venom of the coastal taipan which differed by only a single residue. Primers were designed from Os SNTX-1 to identify related neurotoxins in the venom gland of other Australian elapid snakes by PCR.…”

“…Os SNTX-1 and Pa-a have previously been shown to be toxic to rats and mice with similar intravenous LD 50 values of 63 and 76 mg/kg, respectively. Zamudio et al [19] reported an inability of native Os SNTX-1 to bind neuronal nAChRs (typical of other short-chain neurotoxins) and a reduced affinity for muscle nAChRs, attributing this decrease in binding efficiency potentially to the substitution of either a phenylalanine or a histidine residue with a glycine residue at position 53 of the precursor (Fig. 1a).…”