Diamphotoxin. The arrow poison of the !Kung-Bushmen

Journal of Biological Chemistry (Impact Factor: 4.57). 11/1983; 258(19):11924-31.
Source: PubMed


We have purified the arrow poison extracted from Diamphidia nigro-ornata pupae by the !Kung Bushmen of Southern Africa, and named it diamphotoxin. The toxin is a single chain polypeptide of Mr = 60,000 with an isoelectric point of pH 9.5. It blocks neuromuscular function and is cardiotoxic and hemolytic, and the minimum lethal dose for mice is 25 pg (less than 0.5 fmol). The toxin binds tightly to cells, permitting the resolution of two distinct phases. Erythrocytes exposed to toxin in the absence of divalent cations show no apparent lesion (phase I). After washing and addition of 1 mM Ca2+, there occurs a rapid efflux of K+ followed by the loss of hemoglobin (phase II). The pH optimum for phase I is pH 6.7 and for phase II pH 8.6. The action of the toxin is noncatalytic, requiring a solution concentration of approximately 65 toxin molecules/cell for hemolysis of sheep erythrocytes under standard conditions. Ca2+ ions induce a conformational change in the free, purified toxin molecule. We propose that this change also occurs in membrane-bound toxin. Hemolysis would result from the formation of channels permitting the diffusion of small cations.

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    ABSTRACT: The Bushmen of the Kalahari Desert in Botswana use the pupae of the beetle Diamphidia nigro-ornata Ståhl to poison their arrows. Sequential aqueous extraction, ammonium sulfate precipitation, ultrafiltration and chromatofocusing have given an apparently homogeneous active protein from these pupae with an approximate mol. wt of 54,000, an isoelectric point of about 8.0 pH and a lethal potency (minimum lethal dose, MLD) between 5 and 20 micrograms/kg (i.p. mouse). Preliminary pharmacological studies on less purified material show that, after a delay, this Diamphidia toxin causes sustained contraction of isolated intestinal smooth muscle. This contraction is not blocked by atropine or mepyramine and, therefore, is not due to release of acetylcholine or histamine. Results on the phrenic nerve - hemidiaphragm preparation demonstrate that in the presence of the toxin, contraction in response to indirect stimulation gradually fails and is accompanied by contracture. Since direct stimulation of the muscle still elicits a contraction, the toxin apparently does not affect the contractile mechanism itself. We conclude that Diamphidia pupae contain a protein toxin that is responsible for its lethality. Although this toxin appears to differ in some properties from the toxins reported by Mebs et al., de la Harpe et al. and Kündig, these protein preparations undoubtedly correspond to each other. We did not find any evidence of the low molecular weight toxic component reported by Mebs et al.
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