Soluble expression, purification and functional identification of the framework XV conotoxins derived from different Conus species

• A novel conotoxin cysteine scaffold, framework XV (-C-C-CC-C-C-C-C-), was first identified from Conus litteratus.
• By fused with thioredoxin and 6 × His tag, six XV-conotoxins were successfully expressed in E. coli.
• Different framework XV conotoxins have distinct biological activities on mice and frogs.
• Because of the indel and point mutations occurred in their mature peptides, all the framework XV conotoxins were converged into two branches.
• Position-specific codon conservation is not only restricted to cysteines, but also to other conserved residues.

The conotoxin cysteine framework XV (-C-C-CC-C-C-C-C-), which was named Lt15a, was firstly identified from the cDNA library of Conus litteratus. After that, 18 new framework XV conotoxin sequences were cloned from nine Conus species. Like other conopeptides, the XV-conotoxins have the conserved signal peptide and propeptide, and there are also some conserved residues in their mature peptide. All the framework XV conotoxins were apparently converged into two branches, because of the indel and point mutations occurred in their mature peptides. By fused with thioredoxin and 6 × His tag, six XV-conotoxins were successfully expressed in Escherichia coli and purified. Different framework XV conotoxins have distinct biological activities on mice and frogs, and that may be related to the diversity of the toxin sequences. All the six XV-conotoxins had no obvious effects on the sodium currents of DRG neuron cells of Sprague–Dawley (SD) rats. The identification of this framework of conotoxins enriches our understanding of the structural and functional diversity of conotoxin.