Evaluating the potential of a loop-extended scorpion toxin-like peptide as a protein scaffold

Grafting of exogenous bioactive sites or functional motifs onto structurally stable scaffolds to gain new functions represents an important research direction in protein engineering. Some engineered proteins have been developed into therapeutic drugs. MeuNaTxα-3 (abbreviated as MT-3) is a newly characterized scorpion sodium channel toxin-like peptide isolated from the venom of the scorpion Mesobuthus eupeus, which contains a rigid scaffold highly similar to classical scorpion sodium channel toxins and an extension of eight amino acids in its J-loop region. This extended loop constitutes a flexible region extruded from the scaffold and could be substituted by exogenous functional sequences. In this study, we experimentally evaluated the scaffold potential of MT-3 through grafting two small antimicrobial motifs to replace residues within the loop. Functional assays showed that the two engineered molecules exhibited elevated antimicrobial potency, as compared with the unmodified scaffold, without structural disruption, providing experimental evidence in favor of MT-3 as a promising scaffold in protein engineering.