Modification of the Bacillus sphaericus 51- and 42-kilodalton mosquitocidal proteins: effects of internal deletions, duplications, and formation of hybrid proteins.

The 51- and 42-kDa proteins which constitute the binary mosquitocidal toxin of Bacillus sphaericus 2362 have a low overall sequence similarity but share several regions of near identity (L. Baumann, A. H. Broadwell, and P. Baumann, J. Bacteriol. 170:2045-2050, 1988). By using site-directed mutagenesis, deletions of 6 to 16 amino acids in three of these regions of the 51- and 42-kDa proteins were made, and the modified proteins were expressed in Bacillus subtilis. Deletions in both of these proteins resulted in a loss of toxicity for mosquito larvae. Hybrid proteins containing exchanged fragments of the 51- and 42-kDa proteins were inactive when tested in a variety of combinations, thereby indicating that potentially analogous fragments of these two proteins were not functionally equivalent. An internal duplication of 73 amino acids in the 51-kDa protein and 72 amino acids in the 42-kDa protein resulted in a major reduction in toxicity. These results indicate that the conserved regions of the 51- and 42-kDa proteins are necessary for toxicity to larvae and that the 51- and 42-kDa proteins, despite their sequence similarity, are unique, differing from each other by at least one essential attribute.