Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1989 Sep;171(9):5141–5147. doi: 10.1128/jb.171.9.5141-5147.1989

Evidence for two different types of insecticidal P2 toxins with dual specificity in Bacillus thuringiensis subspecies.

C N Nicholls 1, W Ahmad 1, D J Ellar 1
PMCID: PMC210328  PMID: 2570060

Abstract

Analysis of polypeptides in the crystalline delta-endotoxins from different Bacillus thuringiensis strains revealed two antigenically similar forms of the P2 protein which differed in molecular mass, peptide profile, and amino acid sequence. Purified preparations of the two forms displayed the characteristic dual toxicity of the P2 protein towards members of the orders Lepidoptera and Diptera in vivo but differed markedly in potency for the insects tested. Both species of the P2 protoxin, solubilized and activated by sequential proteolysis with insect gut extract and alpha-chymotrypsin, retained activity in vivo and in vitro, despite the removal of 144 residues from the N terminus. For the low-molecular-mass form, the dual insecticidal activity was reproducible in the in vitro assays.

Full text

PDF
5142

Images in this article

5143Image
on p.5143
5143Image
on p.5143
5144Image
on p.5144

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Adang M. J., Staver M. J., Rocheleau T. A., Leighton J., Barker R. F., Thompson D. V. Characterized full-length and truncated plasmid clones of the crystal protein of Bacillus thuringiensis subsp. kurstaki HD-73 and their toxicity to Manduca sexta. Gene. 1985;36(3):289–300. doi: 10.1016/0378-1119(85)90184-2. [DOI] [PubMed] [Google Scholar]
  2. Bulla L. A., Jr, Bechtel D. B., Kramer K. J., Shethna Y. I., Aronson A. I., Fitz-James P. C. Ultrastructure, physiology, and biochemistry of Bacillus thuringiensis. Crit Rev Microbiol. 1980;8(2):147–204. doi: 10.3109/10408418009081124. [DOI] [PubMed] [Google Scholar]
  3. Burges H. D., Hillyer S., Chanter D. O. Effect of ultraviolet and gamma rays on the activity of delta-endotoxin protein crystals of Bacillus thuringiensis. J Invertebr Pathol. 1975 Jan;25(1):5–9. doi: 10.1016/0022-2011(75)90279-7. [DOI] [PubMed] [Google Scholar]
  4. Cleveland D. W., Fischer S. G., Kirschner M. W., Laemmli U. K. Peptide mapping by limited proteolysis in sodium dodecyl sulfate and analysis by gel electrophoresis. J Biol Chem. 1977 Feb 10;252(3):1102–1106. [PubMed] [Google Scholar]
  5. Donovan W. P., Dankocsik C. C., Gilbert M. P., Gawron-Burke M. C., Groat R. G., Carlton B. C. Amino acid sequence and entomocidal activity of the P2 crystal protein. An insect toxin from Bacillus thuringiensis var. kurstaki. J Biol Chem. 1988 Jan 5;263(1):561–567. [PubMed] [Google Scholar]
  6. Granum P. E., Pinnavaia S. M., Ellar D. J. Comparison of the in vivo and in vitro activity of the delta-endotoxin of Bacillus thuringiensis var. morrisoni (HD-12) and two of its constituent proteins after cloning and expression in Escherichia coli. Eur J Biochem. 1988 Mar 15;172(3):731–738. doi: 10.1111/j.1432-1033.1988.tb13950.x. [DOI] [PubMed] [Google Scholar]
  7. Haider M. Z., Ellar D. J. Analysis of the molecular basis of insecticidal specificity of Bacillus thuringiensis crystal delta-endotoxin. Biochem J. 1987 Nov 15;248(1):197–201. doi: 10.1042/bj2480197. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Haider M. Z., Knowles B. H., Ellar D. J. Specificity of Bacillus thuringiensis var. colmeri insecticidal delta-endotoxin is determined by differential proteolytic processing of the protoxin by larval gut proteases. Eur J Biochem. 1986 May 2;156(3):531–540. doi: 10.1111/j.1432-1033.1986.tb09612.x. [DOI] [PubMed] [Google Scholar]
  9. Hawkes R., Niday E., Gordon J. A dot-immunobinding assay for monoclonal and other antibodies. Anal Biochem. 1982 Jan 1;119(1):142–147. doi: 10.1016/0003-2697(82)90677-7. [DOI] [PubMed] [Google Scholar]
  10. Hofmann C., Vanderbruggen H., Höfte H., Van Rie J., Jansens S., Van Mellaert H. Specificity of Bacillus thuringiensis delta-endotoxins is correlated with the presence of high-affinity binding sites in the brush border membrane of target insect midguts. Proc Natl Acad Sci U S A. 1988 Nov;85(21):7844–7848. doi: 10.1073/pnas.85.21.7844. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Knowles B. H., Ellar D. J. Characterization and partial purification of a plasma membrane receptor for Bacillus thuringiensis var. kurstaki lepidopteran-specific delta-endotoxin. J Cell Sci. 1986 Jul;83:89–101. doi: 10.1242/jcs.83.1.89. [DOI] [PubMed] [Google Scholar]
  12. Knowles B. H., Ellar D. J. Differential specificity of two insecticidal toxins from Bacillus thuringiensis var. aizawai. Mol Microbiol. 1988 Jan;2(1):153–157. doi: 10.1111/j.1365-2958.1988.tb00016.x. [DOI] [PubMed] [Google Scholar]
  13. Knowles B. H., Francis P. H., Ellar D. J. Structurally related Bacillus thuringiensis delta-endotoxins display major differences in insecticidal activity in vivo and in vitro. J Cell Sci. 1986 Aug;84:221–236. doi: 10.1242/jcs.84.1.221. [DOI] [PubMed] [Google Scholar]
  14. Knowles B. H., Thomas W. E., Ellar D. J. Lectin-like binding of Bacillus thuringiensis var. kurstaki lepidopteran-specific toxin is an initial step in insecticidal action. FEBS Lett. 1984 Mar 26;168(2):197–202. doi: 10.1016/0014-5793(84)80245-8. [DOI] [PubMed] [Google Scholar]
  15. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  16. Laemmli U. K., Favre M. Maturation of the head of bacteriophage T4. I. DNA packaging events. J Mol Biol. 1973 Nov 15;80(4):575–599. doi: 10.1016/0022-2836(73)90198-8. [DOI] [PubMed] [Google Scholar]
  17. Matsudaira P. Sequence from picomole quantities of proteins electroblotted onto polyvinylidene difluoride membranes. J Biol Chem. 1987 Jul 25;262(21):10035–10038. [PubMed] [Google Scholar]
  18. Schnepf H. E., Wong H. C., Whiteley H. R. The amino acid sequence of a crystal protein from Bacillus thuringiensis deduced from the DNA base sequence. J Biol Chem. 1985 May 25;260(10):6264–6272. [PubMed] [Google Scholar]
  19. Shibano Y., Yamagata A., Nakamura N., Iizuka T., Sugisaki H., Takanami M. Nucleotide sequence coding for the insecticidal fragment of the Bacillus thuringiensis crystal protein. Gene. 1985;34(2-3):243–251. doi: 10.1016/0378-1119(85)90133-7. [DOI] [PubMed] [Google Scholar]
  20. Stewart G. S., Ellar D. J. Precursor processing during the maturation of a spore-coat protein in Bacillus megaterium KM. Biochem J. 1983 Feb 15;210(2):411–417. doi: 10.1042/bj2100411. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Stewart G. S., Johnstone K., Hagelberg E., Ellar D. J. Commitment of bacterial spores to germinate. A measure of the trigger reaction. Biochem J. 1981 Jul 15;198(1):101–106. doi: 10.1042/bj1980101. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Thomas W. E., Ellar D. J. Bacillus thuringiensis var israelensis crystal delta-endotoxin: effects on insect and mammalian cells in vitro and in vivo. J Cell Sci. 1983 Mar;60:181–197. doi: 10.1242/jcs.60.1.181. [DOI] [PubMed] [Google Scholar]
  23. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Tyrell D. J., Davidson L. I., Bulla L. A., Jr, Ramoska W. A. Toxicity of parasporal crystals of Bacillus thuringiensis subsp. israelensis to mosquitoes. Appl Environ Microbiol. 1979 Oct;38(4):656–658. doi: 10.1128/aem.38.4.656-658.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Widner W. R., Whiteley H. R. Two highly related insecticidal crystal proteins of Bacillus thuringiensis subsp. kurstaki possess different host range specificities. J Bacteriol. 1989 Feb;171(2):965–974. doi: 10.1128/jb.171.2.965-974.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Yamamoto T., Iizuka T. Two types of entomocidal toxins in the parasporal crystals of Bacillus thuringiensis kurstaki. Arch Biochem Biophys. 1983 Nov;227(1):233–241. doi: 10.1016/0003-9861(83)90366-1. [DOI] [PubMed] [Google Scholar]
  27. Yamamoto T., McLaughlin R. E. Isolation of a protein from the parasporal crystal of Bacillus thuringiensis var. Kurstaki toxic to the mosquito larva, Aedes taeniorhynchus. Biochem Biophys Res Commun. 1981 Nov 30;103(2):414–421. doi: 10.1016/0006-291x(81)90468-x. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES