Wael Gad, Khadija Wahni, Rahma Ben-Abderrazek, Joris Messens, Serge Muyldermans, Didier Vertommen, Balkiss Bouhaouala-Zahar, Department of Bio-engineering Sciences, Structural Biology Brussels, Cellular and Molecular Immunology, Ultrastructure, Brussels Center for Redox Biology, 1050 Brussels, Belgium, Brussels Center for Redox Biology, Structural Biology Research Center, VIB, 1050 Brussels, Belgium, VIB-VUB Center for Structural Biology [Bruxelles], VIB [Belgium]-VIB [Belgium], Structural Biology Brussels (SBB), Vrije Universiteit Brussel (VUB), Laboratoire des Venins et Biomolécules Thérapeutiques - Laboratory of Venoms and Therapeutic Biomolecules (LR11IPT08), Institut Pasteur de Tunis, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP), de Duve Institute, Université Catholique de Louvain = Catholic University of Louvain (UCL), Cellular and Molecular Immunology, Vrije Universiteit Brussel, 1050 Brussels, Belgium, Université de Tunis El Manar (UTM), This work was made possible by financial support from Vlaams Instituute Voor Biotechnologie (VIB) and the Flanders Hercules Foundation [grant number HERC16]. J.M. and S.M. are group leaders at the VIB.W.G. is supported by the European Student ExchangeProgramme ‘Erasmus Mundus ECW II’ [grant number 141085-EM-12008-BE-ERA Mundus – ECW]. D.V. is ‘collaborateur logistique of the FNRS-FRS Belgium’ R.B.A. is a Post-Doc in the group of B.B.Z. B.B.Z. is ‘scorpion venoms and antivenoms’ group leader at the IPT, and UCL - SSS/DDUV - Institut de Duve
Envenoming following scorpion sting is a common emergency in many parts of the world. During scorpion envenoming, highly toxic small polypeptides of the venom diffuse rapidly within the victim causing serious medical problems. The exploration of toxin structure-function relationship would benefit from the generation of soluble recombinant scorpion toxins in Escherichia coli. We developed an in vitro wheat germ translation system for the expression of the highly toxic Aah (Androctonus australis hector)II protein that requires the proper formation of four disulphide bonds. Soluble, recombinant N-terminal GST (glutathione S-transferase)-tagged AahII toxin is obtained in this in vitro translation system. After proteolytic removal of the GST-tag, purified rAahII (recombinant AahII) toxin, which contains two extra amino acids at its N terminal relative to the native AahII, is highly toxic after i.c.v. (intracerebroventricular) injection in Swiss mice. An LD50 (median lethal dose)-value of 10 ng (or 1.33 pmol), close to that of the native toxin (LD50 of 3 ng) indicates that the wheat germ in vitro translation system produces properly folded and biological active rAahII. In addition, NbAahII10 (Androctonus australis hector nanobody 10), a camel single domain antibody fragment, raised against the native AahII toxin, recognizes its cognate conformational epitope on the recombinant toxin and neutralizes the toxicity of purified rAahII upon injection in mice., A wheat germ embryo derived cell-free translation system expresses a biologically active, highly toxic scorpion venom protein that is fully neutralized by a camel single domain antibody fragment raised against the native scorpion toxin.