A NOVEL NEUROTOXIN, COBROTOXIN-B, FROM NAJA-NAJA-ATRA (TAIWAN COBRA) VENOM - PURIFICATION, CHARACTERIZATION, AND GENE ORGANIZATION

A novel neurotoxin, cobrotoxin b, was isolated from Naja naja atra (Ta iwan cobra) venom by successive chromatographies on gel filtration and SP-Sephadex C-25 columns. The yield of this novel toxin was 5% of tha t of cobrotoxin from the same venom. Its neurotoxicity determined as t he inhibition of acetylcholine-induced muscle contractions was approxi mately 50% of that of cobrotoxin. Cobrotoxin b consists of 61 amino ac id residues including 8 cysteine residues. Moreover, there are 12 amin o acid substitutions between cobrotoxin b and cobrotoxin. The genomic DNA, with a size of 2,386 bp, encoding the precursor of cobrotoxin b w as isolated from the liver of N. naja atra. The gene consists of three exons separated by two introns. This exon/intron structure is essenti ally the same as that reported for the cobrotoxin gene. Moreover, the nucleotide sequences of the two neurotoxin genes exhibit 92% identity. These results highly suggest that the cobrotoxin b and cobrotoxin gen es are derived from a common ancestor. Comparative analyses of cobroto xin b and cobrotoxin precursors showed that the protein-coding regions of the exons are more diverse than introns, except for in the signal peptide domain. This indicates that the protein-coding regions may hav e arised via accelerated evolution. BLAST searches for sequence simila rity in the GeneBank databases showed that intron 1 of the cobrotoxin b and cobrotoxin genes encodes a small nucleolar RNA (snoRNA). However , the snoRNA gene is absent from the gene encoding the Laticauda semif asciata erabutoxin c precursor (L. semifasciata and N. naja atra are s ea and land snakes, respectively). Since previous studies suggested th e potential mobility of snoRNA genes during evolution, we propose that intron insertions or deletions of snoRNA genes occurred with the evol utionary divergence between the sea snake and land snake neurotoxins.