Cloning, characterization and phylogenetic analyses of members of three major venom families from a single specimen of Walterinnesia aegyptia.

ABSTRACT

Walterinnesia aegyptia is a monotypic elapid snake inhabiting in Africa and Mideast. Although its envenoming is known to cause rapid deaths and paralysis, structural data of its venom proteins are rather limited. Using gel filtration and reverse-phase HPLC, phospholipases A(2) (PLAs), three-fingered toxins (3FTxs), and Kunitz-type protease inhibitors (KIns) were purified from the venom of a single specimen of this species caught in northern Egypt. In addition, specific primers were designed and PCR was carried out to amplify the cDNAs encoding members of the three venom families, respectively, using total cDNA prepared from its venom glands. Complete amino acid sequences of two acidic PLAs, three short chain 3FTxs, and four KIns of this venom species were thus deduced after their cDNAs were cloned and sequenced. They are all novel sequences and match the mass data of purified proteins. For members of each toxin family, protein sequences were aligned and subjected to molecular phylogenetic analyses. The results indicated that the PLAs and a Kunitz inhibitor of W. aegyptia are most similar to those of king cobra venom, and its 3FTxs belongs to either Type I alpha-neurotoxins or weak toxins of orphan-II subtype. It is remarkable that both king cobra and W. aegyptia cause rapid deaths of the victims, and a close evolutionary relationship between them is speculated.