δ-Atracotoxins from Australian Funnel-web Spiders Compete with Scorpion α-Toxin Binding but Differentially Modulate Alkaloid Toxin Activation of Voltage-gated Sodium Channels

doi:10.1074/jbc.273.42.27076

δ-Atracotoxins from Australian Funnel-web Spiders Compete with Scorpion α-Toxin Binding but Differentially Modulate Alkaloid Toxin Activation of Voltage-gated Sodium Channels*

From the ^‡Department of Health Sciences, University of Technology, Sydney, Broadway, New South Wales 2007, Australia, the ^¶Department of Pharmacology, University of Sydney, New South Wales 2006, Australia, ^‖Deakin Research Ltd., CSIRO Division of Food Processing, North Ryde, New South Wales 2113, Australia, ^‡UMR 6560 CNRS, Laboratoire de Biochimie, Université de la Mediterranée, I. F. R. Jean Roche, 13916 Marseille, Cedex 20, France, and ^§CEA, C. E. Saclay, Département d’Ingénierie et d’Etudes des Protéines, Gif-sur-Yvette F-91911, France

Abstract

δ-Atracotoxins from the venom of Australian funnel-web spiders are a unique group of peptide toxins that slow sodium current inactivation in a manner similar to scorpion α-toxins. To analyze their interaction with known sodium channel neurotoxin receptor sites, we studied their effect on [³H]batrachotoxin and ¹²⁵I-Lqh II (where Lqh is α-toxin II from the venom of the scorpion Leiurus quinquestriatus hebraeus) binding and on alkaloid toxin-stimulated ²²Na⁺ uptake in rat brain synaptosomes. δ-Atracotoxins significantly increased [³H]batrachotoxin binding yet decreased maximal batrachotoxin-activated ²²Na⁺ uptake by 70–80%, the latter in marked contrast to the effect of scorpion α-toxins. Unlike the inhibition of batrachotoxin-activated²²Na⁺ uptake, δ-atracotoxins increased veratridine-stimulated ²²Na⁺ uptake by converting veratridine from a partial to a full agonist, analogous to scorpion α-toxins. Hence, δ-atracotoxins are able to differentiate between the open state of the sodium channel stabilized by batrachotoxin and veratridine and suggest a distinct sub-conductance state stabilized by δ-atracotoxins. Despite these actions, low concentrations of δ-atracotoxins completely inhibited the binding of the scorpion α-toxin, ¹²⁵I-Lqh II, indicating that they bind to similar, or partially overlapping, receptor sites. The apparent uncoupling between the increase in binding but inhibition of the effect of batrachotoxin induced by δ-atracotoxins suggests that the binding and action of certain alkaloid toxins may represent at least two distinguishable steps. These results further contribute to the understanding of the complex dynamic interactions between neurotoxin receptor site areas related to sodium channel gating.

Footnotes

↵* This work was supported in part by an Australian Research Council research grant and a University of Technology, Sydney internal research grant (to G. M. N.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵** Present address: The Australian Proteome Analysis Facility, Macquarie University, North Ryde, New South Wales 2109, Australia.
↵§§ To whom correspondence should be addressed: CEA, C.E.-Saclay, Département d’Ingénierie et d’Etudes des Protéines, Gif-sur-Yvette, F-91911, France. Fax: 33 1 69 08 90 71; E-mail:GORDON{at}dsvidf.cea.fr.
↵¶¶ To whom correspondence should be addressed: Dept. of Health Sciences, University of Technology, Sydney, P. O. Box 123, Broadway, New South Wales 2007, Australia. Fax: 61 2 9514–2228; E-mail: Graham.Nicholson{at}uts.edu.au.
Abbreviations:

δ-atracotoxin-Ar1

(formerly robustoxin) from the venom of the spiderAtrax robustus

δ-atracotoxin-Hv1

(formerly versutoxin) from the venom of the spider Hadronyche versuta

TTX

tetrodotoxin

AaH II

α-toxin II from the venom of the scorpionAndroctonus australis hector

ATX II

sea anemone toxin II from Anemonia sulcata

[³H]BTX

[³H]batrachotoxinin A-20α-benzoate

Lqh II

α-toxin II from the venom of the scorpion Leiurus quinquestriatus hebraeus

TEA

tetraethylammonium

HPLC

high performance liquid chromatography

Tricine

N-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]-glycine

BSA

bovine serum albumin.
- Received June 22, 1998.
- Revision received August 1, 1998.