Definition of a saxitoxin (STX) binding code enables discovery and characterization of the anuran saxiphilin family

American bullfrog (Rana castesbeiana) saxiphilin (RcSxph) is a high-affinity toxin sponge protein thought to prevent intoxication by saxitoxin (STX), a lethal bisguanidinium neurotoxin that causes paralytic shellfish poisoning (PSP) by blocking voltage-gated sodium channels (NaVs). How specific RcSxph interactions contribute to STX binding has not been defined and whether other organisms have similar proteins is unclear. Here, we use mutagenesis, ligand binding, and structural studies to define the energetic basis of Sxph:STX recognition. The resultant STX recognition code enabled engineering of RcSxph to improve its ability to rescue NaVs from STX and facilitated discovery of 10 new frog and toad Sxphs. Definition of the STX binding code and Sxph family expansion among diverse anurans separated by similar to 140 My of evolution provides a molecular basis for understanding the roles of toxin sponge proteins in toxin resistance and for developing novel proteins to sense or neutralize STX and related PSP toxins.

Automated summary

This study defines the energetic basis of recognition between the sponge protein, RcSxph, and the neurotoxin STX. The study also improves the ability of RcSxph to neutralize STX and discovers new frog and toad sponge proteins. This research provides a molecular basis for understanding the roles of toxin sponge proteins in toxin resistance and developing novel proteins to sense or neutralize STX and related PSP toxins.