Studies on the delta-endotoxin ofBacillus thuringiensisvar.tenebrionis

Abstract

Parasporal crystals of the recently isolated Bacillus thuringiensis var. tenebrionis are toxic for coleopteran larvae. Unlike those of other strains they are soluble either in aqueous solutions of NaBr at neutral pH or in water after titration to pH values above pH 10.0. The dissolved crystal protein readily forms crystals after removal of the salt or neutralization. The crystal protein was not found to differ much in the amino acid composition from other crystal proteins. The parasporal crystals are composed of subunits of Mr 68 000 which are not linked by disulfide bridges.

Proteolytic processing of a coleopteran-specific δ-endotoxin produced by Bacillus thuringiensis var. tenebrionis

“…tenebrionis 4-endotoxin crystals have been previously reported to contain polypeptides of 74 and 68 kDa (Krieg et al, 1987) or 68 and 55 kDa (Bernhard, 1986). Our data indicate that this discrepancy could have resulted from harvesting the crystals at different times after sporulation.…”

“…The observation that, at stage VI, when crystal formation is apparently complete, the major polypeptide present is 73 kDa (Figs. 4a and 4b ) leads to the suggestion that proteolysis to the 67 kDa (T3a) polypeptide is the result of associated endogenous proteinase activity (Bernhard, 1986) after crystal release. This increase in T 3a during crystal release could be effectively inhibited by using proteinase inhibitors (Figs.…”

“…After the isolation of a coleopteran-specific 8-endotoxin (Krieg et al, 1983;Herrnstadt et al, 1986), the initial analysis of the crystalline toxin inclusions by SDS/polyacrylamide-gel electrophoresis (Bernhard, 1986;Herrnstadt et al, 1986) revealed a major polypeptide of 68 or 64 kDa. No toxic polypeptides smaller than 64 kDa were obtained when this protein was treated with trypsin or gut extracts from a susceptible larva (Herrnstadt et al, 1986).…”

Analytical Chemistry of Bacillus thuringiensis

“…The latter process can be accomplished with salt solutions, high pH, low pH, urea or surfactants such as sodium dodecyl sulfate (SDS) (15,22). As mentioned previously, the BT subspecies tenebrionus or san diego protoxins lack the disulfide bonds and can be solubilized with 3 M sodium bromide solution without reductant (10).…”

“…The smaller proteins, such as those of subspecies tenebrionus or san diego. are held together by hydrophobic and electrostatic interactions which can be overcome in concentrated salt solutions (10). All of the active protein toxins of BT act by binding to the insect's midgut epithelium and disrupting the integrity of membranes.…”