Specificity-determining regions of a lepidopteran-specific insecticidal protein produced by Bacillus thuringiensis

• Published in: The Journal of biological chemistry Vol. 265; no. 34; pp. 20923 - 20930
• Main Authors: Schnepf, H E, Tomczak, K, Ortega, J P, Whiteley, H R
• Format: Journal Article
• Language: English
• Published: Bethesda, MD Elsevier Inc 05.12.1990; American Society for Biochemistry and Molecular Biology
• Subjects: Amino Acid Sequence; AMINO ACID SEQUENCES; Animals; BACILLUS THURINGIENSIS; Bacillus thuringiensis - genetics; Bacillus thuringiensis Toxins; BACTERIA; Bacterial Proteins - genetics; Bacterial Proteins - pharmacology; Bacterial Toxins - genetics; Bacteriology; Biological and medical sciences; Chimera; codons; CryIA(a) protein; CryIA(c) protein; Endotoxins; Fundamental and applied biological sciences. Psychology; GENE; GENES; Genes, Bacterial; HELIOTHIS VIRESCENS; Hemolysin Proteins; INSECTICIDAL PROPERTIES; INSECTICIDAS; INSECTICIDE; INSECTICIDES; Larva; Lepidoptera; Lepidoptera - drug effects; MANDUCA SEXTA; Microbiology; Molecular Sequence Data; Moths - drug effects; Mutagenesis, Site-Directed; Noctuidae; NUCLEOTIDE; NUCLEOTIDES; NUCLEOTIDOS; Pathogenicity, virulence, toxins, bacteriocins, pyrogens, host-bacteria relations, miscellaneous strains; Plasmids; PROTEINAS UNICELULARES; PROTEINE MICROBIOLOGIQUE; Restriction Mapping; SINGLE CELL PROTEINS; Sphingidae; Heliothis virescens; Insecticide; Insecta; Bacillus thuringiensis; Molecular hybrid; Cytotoxicity; Bacillaceae; Sphingidae; Proteins; Bacillales; Gene; Structure activity relation; Arthropoda; Lepidoptera; Manduca sexta; Bacteria; Noctuidae; Invertebrata
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(17)45305-1

The lepidopteran-specific, insecticidal crystal proteins of Bacillus thuringiensis vary in toxicity to different species of lepidopteran larvae. We report studies of CryIA(a) and CryIA(c), two related proteins that have different degrees of toxicity to Heliothis virescens yet very similar degrees of toxicity to Manduca sexta. The amino acid differences between these proteins are located primarily between residues 280 and 722. We have constructed a series of chimeric proteins and determined their toxicities to both insects. The most significant findings arise from the replacement of three segments of the cryIA(c) gene with homologous portions of the cryIA(a) gene: codons 332-428, 429-447, and 448-722. Each of these segments contributed substantially and largely additively toward efficacy for H. virescens. However, replacement of the 429-447 segment of cryIA(c) gene with the cryIA(a) sequence resulted in a 27-50-fold reduction in toxicity toward M. sexta whereas the reduction in toxicity to H. virescens was only 3-4-fold. Subdivision of the 429-447 segment and replacements involving residues within this segment reduced toxicity to M. sexta by 5- to more than 2000-fold whereas toxicity to H. virescens was only reduced 3-10-fold. These observations indicate that: 1) different but overlapping regions of the cryIA(c) gene determine specificity to each of the two test insects; 2) some of the examined gene segments interact in determining specificity; and 3) different sequences in the cryIA(a) and cryIA(c) genes are required for maximal toxicity to M. sexta.