Isolation, molecular cloning and functional characterization of a novel beta-toxin from the Venezuelan scorpion, Tityus zulianus

Sting in children by Tityus zulianus scorpions (western Venezuela) often produces cardiorespiratory arrest and death by pulmonary oedema. To assess its toxicity, lethality in mice of T. zulianus soluble venom was determined. Toxin composition was studied by fractionating the crude venom through reversed-phase HPLC. The most abundant peptide, Tz1, was purified further and its N-terminal sequence, amino acid composition and molecular mass (by electron-spray ionization mass spectrometry) determined. In the presence of Tz1, activation of recombinant rat skeletal muscle sodium channels (Na(V)1.4) was shifted about 35 mV in the hyperpolarizing direction in a prepulse-dependent manner. This typical beta-toxin effect had an apparent EC50 of 3.5 microM A cDNA sequence encoding Tz1 was isolated from T. zulianus venom gland RNA using a combination of 5'- and 3'-RACE PCR. Analysis of the encoded sequence indicated that Tz1 is the processed product of a precursor containing: (i) a 20-residue long leader peptide; (ii) the amino acid sequence of the mature toxin (64 residues); and (iii) an extra Gly-Lys tail at the C-terminus, probably removed post-translationally. A comparison of Tz1 with Tityus serrulatus beta-toxin Ts1 revealed that some of the non-conservative replacements in Tz1 lie in regions potentially involved in receptor recognition.