PRODUCTION OF MULTIPLE DELTA-ENDOTOXINS BY BACILLUS-THURINGIENSIS - DELTA-ENDOTOXINS PRODUCED BY STRAINS OF THE SUBSPECIES GALLERIAE AND WUHANENSIS

A method was developed to assess the number of delta-endotoxins contained in Bacillus thuringiensis entomocidal crystals. It utilized proteolytic conversion of 13O-kDa protoxin into 60- to 65-kDa ''true'' toxin via limited proteolysis with trypsin and separation of stable N-terminal domains by fast-performance liquid chromatography. Immunodiffusion experiments and N-terminal sequence determination (applied to the major component isolated by SDS-PAGE) completed the analysis of the crystal protein composition. The application of this approach to crystals produced by cells of B. thuringiensis subsp. galleriae and wuhanensis allowed us to identify at least seven and eight different delta-endotoxins, respectively. Among those delta-endotoxins assigned to previously described families, CryIA, CryID, CryIF, and CryIG were found, as well as crystal proteins, which possess N-terminal amino acid sequences very different from those of all known delta-endotoxins. Possible functional consequences of delta-endotoxin multiplicity are discussed.