Construction by site-directed mutagenesis of a 39-kilodalton mosquitocidal protein similar to the larva-processed toxin of Bacillus sphaericus 2362.
After ingestion of the parasporal crystals of Bacillus sphaericus, mosquito larvae process the 42-kilodalton (kDa) toxin to a protein of 39 kDa, which has an increased toxicity (A. H. Broadwell and P. Baumann, Appl. Environ. Microbiol. 53:1333-1337, 1987). A similar activation is performed by trypsin and chymotrypsin. Using site-directed mutagenesis, we have constructed derivatives of the 42-kDa toxin with a deletion of 10 amino acids at the N terminus and deletions of 7, 17, or 20 amino acids at the C terminus. Toxicity for mosquito larvae was retained upon deletion of 7 or 17 amino acids but was lost upon deletion of 20 amino acids. Evidence is presented indicating that the protein containing deletions of 10 amino acids at the N terminus and 17 amino acids at the C terminus (corresponding to potential chymotrypsin cleavage sites) is similar to the 39-kDa protein produced in mosquito larvae or by digestion with chymotrypsin. Digestion with trypsin appears to generate a protein lacking 16 or 19 amino acids from the N terminus and 7 amino acids from the C terminus. As is the case with the recombinant-made 42-kDa protein, toxicity of its derivatives is dependent on the presence of a 51-kDa protein which is a component of the parasporal crystal of B. sphaericus 2362.
Broadwell
A H
AH
Department of Microbiology, University of California, Davis 95616.
Clark
M A
MA
Baumann
L
L
Baumann
P
P
D000595:Amino Acid Sequence; D000818:Animals; D001407:Bacillus / Q000235:genetics*; D001412:Bacillus subtilis / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics* / Q000633:toxicity; D001427:Bacterial Toxins / Q000235:genetics*; D001483:Base Sequence; D003465:Culex / Q000187:drug effects; D015870:Gene Expression; D007814:Larva; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009154:Mutation; D015345:Oligonucleotide Probes; D015183:Restriction Mapping
D001426:Bacterial Proteins; D001427:Bacterial Toxins; D015345:Oligonucleotide Probes; C046134:binB protein, Bacillus sphaericus