The successful synthesis and expression of functionally active recombinant charybdotoxin provides a tool for studying the fundamental mechanisms of toxin-K+ channel interactions.
Recombinant CTX enables scalable toxin production; leaves open its utility as a tool for cardiac K+ channel studies.
A gene encoding charybdotoxin (CTX), a K+ channel blocker from scorpion venom, was designed, synthesized, and expressed as a cleavable fusion protein in Escherichia coli. A sequence-specific protease, factor Xa, was used to cleave the fusion protein and thus release the toxin peptide. The recombinant toxin was purified, oxidized to form disulfide bonds, and treated to form N-terminal pyroglutamate. Recombinant CTX is identical to the native venom CTX with respect to high-performance liquid chromatography mobility, amino acid composition, and N-terminal modification. With single Ca2(+)-activated K+ channels as an assay system, recombinant CTX shows blocking and dissociation kinetics identical to the native venom toxin. The synthetic gene and high-level expression of functionally active CTX make it possible to study the fundamental mechanism of the toxin-ion channel interaction.
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Park et al. (1991) studied this question.
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