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

• Published in: Toxicon (Oxford) Vol. 43; no. 6; pp. 671 - 684
• Main Authors: Borges, Adolfo, Alfonzo, Marcelo J., Garcı́a, Carmen C., Winand, Nena J., Leipold, Enrico, Heinemann, Stefan H.
• Format: Journal Article
• Language: English
• Published: Oxford Elsevier Ltd 01.05.2004; Elsevier Science
• Subjects: Animal poisons toxicology. Antivenoms; Animals; Base Sequence; Beta-scorpion toxin; Biological and medical sciences; Cloning, Molecular; DNA Primers; Medical sciences; Mice; Molecular Sequence Data; Neurotoxins - genetics; Neurotoxins - toxicity; Receptor site-4; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger - analysis; Scorpion Venoms - genetics; Scorpion Venoms - toxicity; Sodium channel activation; Tityus; Tityus zulianus; Toxicology; Venezuela; K V, voltage-gated potassium channel; Beta-scorpion toxin; RACE, random amplification of cDNA ends; Receptor site-4; ESI/MS, electrospray ionization mass spectrometry; Tityus zulianus; Sodium channel activation; Tityus; Na V, voltage-gated sodium channel; TFA, trifluoroacetic acid; Characterization; Toxin; Arachnida; Scorpionides; Arthropoda; Animal origin; Isolation; Invertebrata; Molecular cloning
• ISSN: 0041-0101; 1879-3150
• DOI: 10.1016/j.toxicon.2004.02.022

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 V1.4) was shifted about 35 mV in the hyperpolarizing direction in a prepulse-dependent manner. This typical β-toxin effect had an apparent EC 50 of 3.5 μM. 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 β-toxin Ts1 revealed that some of the non-conservative replacements in Tz1 lie in regions potentially involved in receptor recognition.