Journal
TOXICON
Volume 39, Issue 5, Pages 693-702Publisher
PERGAMON-ELSEVIER SCIENCE LTD
DOI: 10.1016/S0041-0101(00)00199-9
Keywords
Anthopleura elegantissima; Carcinus maenas; polypeptide toxins; neurotoxicity; cardiotoxicity
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Five toxins (APE 1 to APE 5) of the sea anemone species Anthopleura elegantissima (Brandt) have been isolated from a toxic by-product fraction of its concentrated crude watery-methanolic extract, prepared previously for the isolation of a neuropeptide (the head-activator) by Schaller and Bodenmuller (Proc. Natl. Acad. Sci. USA 78 (1981) 7000) from 200 kg sea anemones. Toxin purification was performed by desalting of the starting material by dialysis (MWCO 3500) against distilled water, anion exchange chromatography on QAE-Sephadex A25 at pH 8, twice gel filtration on Sephadex G50 m, repeated chromatography on QAE-Sephadex at pH 10 and chromatography on the cation exchanger Fractogel(R) EMD SO3--650 M. Final purification of the toxins was achieved by HPLC on MN SP 250/10 Nucleosil(R) 500-5 C-18 PPN and MN SP 250/21 Nucleosil(R) 300-7 C-18. Each toxin was composed of at least two isotoxins of which APE 1-1, APE 1-2, APE 2-1, APE 2-2 and APE 5-3 were isolated in preparative scale. With exception of APE 5-3 the sequences of the isotoxins have been elucidated. They resemble the 47 residue type-I long polypeptide toxins native to Anemonia sulcata (Pennant). All isotoxins paralyse the shore crab (Carcinus maenas) by tetanic contractions after i.m. application. The toxins modify current passing through the fast Na+ channel in neuroblastoma cells, leading to delayed and incomplete inactivation. APE 1-1, APE 2-1 and APE 5-3 produce a positive inotropic effect in mammalian heart muscle, although they differ in potency. The order of potency is APE 2-1 > APE 1-1 > APE 5-3 (i.e, threshold concentrations are 1, 10 and 300 nM, respectively). In addition, they enhance the spontaneous beating frequency in isolated right atria (guinea pig). The most potent cardiotoxic isotoxin is APE 2-1, its sequence is identical with that of AP-C, a toxin isolated and characterised previously by Norton ct al. (Drugs and Feuds from the Sea, 1978, University of Oklahoma Press, p. 37-50). LD50 APE 2-1: 1 mug/kg b,.w. C. maenas (i.m.) LD50 APE 1-1: 10 mug/kg b.w. C. maenas (i.m.) LD50 APE 5-3: 50 mug/kg, b.w. C. maenas (i.m.) (C) 2000 Elsevier Science Ltd. All rights reserved.
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