Lethal toxins and cross-neutralization of venoms from the African water cobras, Boulengerina annulata annulata and Boulengerina christyi

Abstract

Venoms of the water cobras, Boulengerina, were assayed for lethality, proteolytic activity and protein content. Boulengerina annulata annulata and B. christyi venoms averaged 89% protein and lacked proteolytic activity. The murine i.p. LD50 of B. a. annulata and B. christyi venoms were 0.143 and 0.120 mg/kg, respectively. Polyvalent antivenom produced by the South African Institute of Medical Research neutralized 575 and 200 LD50 of B. a. annulata and B. christyi venoms/ml antivenom, respectively. Cation exchange chromatography resolved four lethal peaks from B. a. annulata venom and six lethal peaks from B. christyi venom. The major lethal peaks (about 12% of total venom protein) were purified further with molecular sieve chromatography and were characterized as 61 (B. a. annulata toxin) and 62 (B. christyi toxin) residue polypeptides with four half-cystines. Elucidation of the complete amino acid sequences indicated that these toxins belonged to the short-chain class of postsynaptic neurotoxins. Short-chain neurotoxins 1 from B. a. annulata and B. christyi had murine i.p. LD50 of 0.052 and 0.083 mg/kg, respectively, and showed over 80% homology with N. nigricollis alpha toxin. Reverse-phase analysis of another peak present in both venoms resolved a toxin that had an N-terminus identical to B. christyi short-chain neurotoxin 1. These fractions also contained toxins readily separable from the short-chain isotoxin by preparative reverse-phase chromatography. Amino acid sequencing of the first 28 residues indicated that both toxins were long-chain neurotoxins with identical N-termini. The LD50 of long-chain neurotoxins 2 from B. a. annulata and B. christyi venoms were 0.086 and 0.090 mg/kg, respectively. The venoms of these little-known elapids have the lowest LD50 of any African proteroglyph studied thus far and have high concentrations of potent postsynaptic neurotoxins.