Solution structure of a short-chain insecticidal toxin LaIT1 from the venom of scorpion Liocheles australasiae.

The solution structure of an insecticidal toxin LaIT1, a 36-residue peptide with a unique amino-acid sequence and two disulfide bonds, isolated from the venom of the scorpion Liocheles australasiae was determined by heteronuclear NMR spectroscopy. Structural similarity search showed that LaIT1 exhibits an inhibitory cystine knot (ICK)-like fold, which usually contains three or more disulfide bonds. Mutational analysis has revealed that two Arg residues of LaIT1, Arg(13) and Arg(15), play significant roles in insecticidal activity.

Improvement of in vitro-transcribed amber suppressor tRNAs toward higher suppression efficiency in wheat germ extract.

In vitro-transcribed, unmodified, and non-aminoacylated amber suppressor tRNAs that are recognized by natural aminoacyl-tRNA synthetase were improved toward higher suppression efficiency in batch-mode cell-free translation in wheat germ extract. The suppression efficiency of the suppressor obtained through four sequence optimization steps (anticodon alteration of natural tRNAs (the first generation); chimerization of the efficient suppressors in the first generation; investigation and optimization of the effective parts in the second generation; combination of the optimized parts in the third generation) and by the terminal tuning was approximately 60%, which was 2.4-fold higher than that of the best suppressor in the first generation. In addition, an eRF1 aptamer further increased the efficiency up to 85%. This highly efficient suppression system also functioned well in a dialysis-based large-scale protein synthesis.

A novel amphipathic linear peptide with both insect toxicity and antimicrobial activity from the venom of the scorpion Isometrus maculatus.

Scorpion venoms are composed of a number of peptides, many of which show neurotoxicity. In addition to these neurotoxins, several antimicrobial peptides have also been isolated from the venoms. The scorpion Isometrus maculatus, belonging to the Buthidae family, is found in many tropical regions including Japan, but little attention has been paid to its biological activity and chemical composition. In this study, we isolated a novel insect toxin, Im-1, by bioassay-guided fractionation of the venom of I. maculatus. Rapid and reversible paralysis was observed after injection of Im-1 into crickets. Im-1 consists of 56 amino acids, and is predicted to form an amphipathic alpha-helix. Since Im-1 shares sequence similarity to an antimicrobial peptide, parabutoporin, we evaluated its effects on several bacterial strains and found that it showed an antimicrobial activity profile similar to parabutoporin. This suggests that Im-1 and parabutoporin exert their antimicrobial effects through similar mechanisms.

Purification and cDNA cloning of LaIT2, a novel insecticidal toxin from venom of the scorpion Liocheles australasiae.

The novel insecticidal toxin, LaIT2, was isolated from venom of the scorpion Liocheles australasiae. The amino acid sequence of LaIT2 was determined by an Edman degradation analysis and subsequent cDNA cloning. LaIT2 is composed of 59 amino acids with three disulfide bridges, and shares sequence similarity to the scorpion beta-KTx peptides.

Purification and characterization of a novel short-chain insecticidal toxin with two disulfide bridges from the venom of the scorpion Liocheles australasiae.

Scorpion venoms contain a variety of peptides toxic to mammals, insects and crustaceans. Most of the scorpion toxins have been isolated from the venoms of scorpions in the family Buthidae, but little interest has been paid to non-Buthidae scorpions. In this study, we isolated a short-chain insecticidal toxin (LaIT1) from the venom of the scorpion Liocheles australasiae belonging to the Hemiscorpiidae family. This toxin showed insect toxicity against crickets at a dose of 1.0 microg/insect, but no toxicity was observed against mice even after injection of 1.0 microg of LaIT1 via the intracerebroventricular route, suggesting that the effect of the toxin is insect-selective. Edman sequencing and mass spectrometric analysis revealed that the toxin is composed of 36 amino acid residues and cross-linked by only two disulfide bridges. The pattern of the disulfide bridges was assigned by LC/MS analysis after enzymatic digestion. LaIT1 shows no sequence homology to any other known toxins, suggesting that this toxin represents a novel structural motif class.

Ladder-shaped polyether compound, desulfated yessotoxin, interacts with membrane-integral alpha-helix peptides.

Ladder-shaped polyether compounds, represented by brevetoxins, ciguatoxins, maitotoxin, and prymnesins, are thought to possess the high affinity to transmembrane proteins. As a model compound of ladder-shaped polyethers, we adopted desulfated yessotoxin (2) and examined its interaction with glycopholin A, a membrane protein known to form a dimer or oligomer. Desulfated yessotoxin turned out to interact with the alpha-helix so as to induce the dissociation of glycopholin oligomers when examined by SDS and PFO gel electrophoresis. The results provided the first evidence that ladder-shaped polyethers interact with transmembrane helix domains.