Functional expression of alpha-latrotoxin in baculovirus system.

To facilitate the study of the mechanism of alpha-latrotoxin action, it is necessary to create a biologically active recombinant toxin. Mature alpha-latrotoxin is naturally produced by post-translational cleavage, probably at two furin sites located at the N- and C-termini of the precursor. A recombinant baculovirus has now been constructed, which encodes the melittin signal peptide fused to the 130-kDa mature toxin between the furin sites. Insect cells, infected with this baculovirus, secreted recombinant alpha-latrotoxin. This was partially purified and proved indistinguishable from the natural toxin with respect to its molecular mass, immunostaining, toxicity to mice, binding to alpha-latrotoxin receptors (latrophilin or neurexin Ialpha) and electrophysiological recording in the mouse diaphragm. The successful expression of recombinant alpha-latrotoxin permits mutational analysis of the toxin.