Cry3A Toxin Research Articles

The oligomeric state of Bacillus thuringiensis Cry toxins in solution

The molecular mass of different Cry toxins produced by Bacillus thuringiensis bacteria was estimated by size-exclusion chromatography and non-denaturing polyacrylamide gel electrophoresis at neutral and alkaline pH in order to assess the existence of oligomers in solution. We found that Cry1Aa, Cry1Ac, Cry1C, Cry1D and Cry3A toxins exist in solution as a mixture of monomer and high molecular mass aggregates with an apparent molecular mass greater than 600 kDa, that depend on the time elapsed between toxin activation and analysis. Aggregation of toxins by disulfide bonds is unlikely because aggregates are also observed in samples incubated with DTT. These data show that the Cry toxins studied do not form oligomers of less than ten subunits in solution and suggest that oligomer formation may occur after the toxin binds to the receptor and inserts into the membrane.

Intramolecular Proteolytic Cleavage ofBacillus thuringiensisCry3A δ-Endotoxin May Facilitate Its Coleopteran Toxicity

The Cry3A δ-endotoxin protein inclusion synthesized byBacillus thuringiensissubsp.tenebrionisis soluble in alkaline and acid buffer solutions but the toxin precipitates when returned to neutral pH conditions. The midgut pH of susceptible beetle larvae is neutral to slightly acidic, a pH environment in which the Cry3A toxin is insoluble. To investigate this paradox we studied the Cry3A toxin after various proteolytic treatments. In many cases the toxin was cleaved into polypeptides that remained associated under nondenaturing conditions. Interestingly a chymotrypsinized Cry3A product was soluble under neutral pH conditions, retained full activity against susceptible beetle larvae, and exhibited specific binding toLeptinotarsa decemlineatamidgut membranes.