A RECOMBINANT ADENOVIRUS THAT DIRECTS SECRETION OF BIOLOGICALLY-ACTIVE KAPPA-BUNGAROTOXIN FROM MAMMALIAN-CELLS

A novel Cre-lox system was used to construct an adenovirus encoding K- bungarotoxin (kappa-Bgt), modified to be secreted by attachment of a b ovine prolactin signal sequence at the N-terminus of the toxin. Wester n blot of medium from HEK-293 cells infected with the virus demonstrat ed that recombinant kappa-Bgt (R-kappa-Bgt) was secreted. The biologic al activity of the secreted R-kappa-Bgt was investigated in Xenopus oo cytes that expressed neuronal nicotinic acetylcholine receptor (nAChR) subtypes alpha 3 beta 2 and alpha 2 beta 2. The recombinant toxin inh ibited the response of alpha 3 beta 2 type AChRs to ACh, but did not i nhibit the response of alpha 2 beta 2 type AChRs. These data demonstra ted that the recombinant adenovirus directs the secretion of biologica lly active kappa-Bgt from a mammalian cell line. Because adenovirus ca n be used to infect post-mitotic cells, recombinant adenoviruses encod ing biologically active peptides may be of use as delivery vehicles fo r in vivo experiments where repeated application of the purified pepti de is unfeasible.