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. 1992 Aug;58(8):2536–2542. doi: 10.1128/aem.58.8.2536-2542.1992

Novel Bacillus thuringiensis insecticidal crystal protein with a silent activity against coleopteran larvae.

B Lambert 1, H Höfte 1, K Annys 1, S Jansens 1, P Soetaert 1, M Peferoen 1
PMCID: PMC195818  PMID: 1514800

Abstract

A novel Bacillus thuringiensis crystal protein with a silent activity against the Colorado potato beetle is described. The crystal proteins are produced as bipyramidal crystals. These crystals contain a protein of 129 kDa with a trypsin-resistant core fragment of 72 kDa. Neither a spore-crystal mixture nor in vitro-solubilized crystals are toxic to any of several Lepidoptera and Coleoptera species tested. In contrast, a trypsin-treated solution containing the 72-kDa tryptic core fragment of the protoxin is highly toxic to Colorado potato beetle larvae. The crystal protein-encoding gene was cloned and sequenced. The inferred amino acid sequence of the putative toxic fragment has 37, 32, and 33% homology to the CryIIIA, CryIIIB, and CryIIID toxins, respectively. Interestingly, the 501 C-terminal amino acids show 41 to 48% amino acid identity with corresponding C-terminal amino acid sequences of other crystal proteins. Because of the toxicity of the fragment to the Colorado potato beetle and because of the distinct similarities of the toxic fragment with the other CryIII proteins, this gene was given a new subclass name (cryIIIC) within the CryIII class of coleopteran-active crystal proteins. CryIIIC represents the first example of a crystal protein with a silent activity towards coleopteran insect larvae. Natural CryIIIC crystals are not toxic. Toxicity is revealed only after an in vitro solubilization and activation step.

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Selected References

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  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. Chungjatupornchai W., Höfte H., Seurinck J., Angsuthanasombat C., Vaeck M. Common features of Bacillus thuringiensis toxins specific for Diptera and Lepidoptera. Eur J Biochem. 1988 Apr 5;173(1):9–16. doi: 10.1111/j.1432-1033.1988.tb13960.x. [DOI] [PubMed] [Google Scholar]
  3. 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]
  4. Donovan W. P., Dankocsik C., Gilbert M. P. Molecular characterization of a gene encoding a 72-kilodalton mosquito-toxic crystal protein from Bacillus thuringiensis subsp. israelensis. J Bacteriol. 1988 Oct;170(10):4732–4738. doi: 10.1128/jb.170.10.4732-4738.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Donovan W. P., Gonzalez J. M., Jr, Gilbert M. P., Dankocsik C. Isolation and characterization of EG2158, a new strain of Bacillus thuringiensis toxic to coleopteran larvae, and nucleotide sequence of the toxin gene. Mol Gen Genet. 1988 Nov;214(3):365–372. doi: 10.1007/BF00330468. [DOI] [PubMed] [Google Scholar]
  6. Herrnstadt C., Gilroy T. E., Sobieski D. A., Bennett B. D., Gaertner F. H. Nucleotide sequence and deduced amino acid sequence of a coleopteran-active delta-endotoxin gene from Bacillus thuringiensis subsp. san diego. Gene. 1987;57(1):37–46. doi: 10.1016/0378-1119(87)90174-0. [DOI] [PubMed] [Google Scholar]
  7. Hindley J., Berry C. Bacillus sphaericus strain 2297: nucleotide sequence of 41.9 kDa toxin gene. Nucleic Acids Res. 1988 May 11;16(9):4168–4168. doi: 10.1093/nar/16.9.4168. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Honée G., van der Salm T., Visser B. Nucleotide sequence of crystal protein gene isolated from B. thuringiensis subspecies entomocidus 60.5 coding for a toxin highly active against Spodoptera species. Nucleic Acids Res. 1988 Jul 11;16(13):6240–6240. doi: 10.1093/nar/16.13.6240. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Höfte H., Seurinck J., Van Houtven A., Vaeck M. Nucleotide sequence of a gene encoding an insecticidal protein of Bacillus thuringiensis var. tenebrionis toxic against Coleoptera. Nucleic Acids Res. 1987 Sep 11;15(17):7183–7183. doi: 10.1093/nar/15.17.7183. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Höfte H., Soetaert P., Jansens S., Peferoen M. Nucleotide sequence and deduced amino acid sequence of new Lepidoptera-specific crystal protein gene from Bacillus thuringiensis. Nucleic Acids Res. 1990 Sep 25;18(18):5545–5545. doi: 10.1093/nar/18.18.5545. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Höfte H., Van Rie J., Jansens S., Van Houtven A., Vanderbruggen H., Vaeck M. Monoclonal Antibody Analysis and Insecticidal Spectrum of Three Types of Lepidopteran-Specific Insecticidal Crystal Proteins of Bacillus thuringiensis. Appl Environ Microbiol. 1988 Aug;54(8):2010–2017. doi: 10.1128/aem.54.8.2010-2017.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Höfte H., Whiteley H. R. Insecticidal crystal proteins of Bacillus thuringiensis. Microbiol Rev. 1989 Jun;53(2):242–255. doi: 10.1128/mr.53.2.242-255.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Höfte H., de Greve H., Seurinck J., Jansens S., Mahillon J., Ampe C., Vandekerckhove J., Vanderbruggen H., van Montagu M., Zabeau M. Structural and functional analysis of a cloned delta endotoxin of Bacillus thuringiensis berliner 1715. Eur J Biochem. 1986 Dec 1;161(2):273–280. doi: 10.1111/j.1432-1033.1986.tb10443.x. [DOI] [PubMed] [Google Scholar]
  14. Lambert B., Theunis W., Aguda R., Van Audenhove K., Decock C., Jansens S., Seurinck J., Peferoen M. Nucleotide sequence of gene cryIIID encoding a novel coleopteran-active crystal protein from strain BTI109P of Bacillus thuringiensis subsp. kurstaki. Gene. 1992 Jan 2;110(1):131–132. doi: 10.1016/0378-1119(92)90457-z. [DOI] [PubMed] [Google Scholar]
  15. MacIntosh S. C., Stone T. B., Sims S. R., Hunst P. L., Greenplate J. T., Marrone P. G., Perlak F. J., Fischhoff D. A., Fuchs R. L. Specificity and efficacy of purified Bacillus thuringiensis proteins against agronomically important insects. J Invertebr Pathol. 1990 Sep;56(2):258–266. doi: 10.1016/0022-2011(90)90109-j. [DOI] [PubMed] [Google Scholar]
  16. Maxam A. M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. doi: 10.1016/s0076-6879(80)65059-9. [DOI] [PubMed] [Google Scholar]
  17. 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]
  18. Sekar V., Thompson D. V., Maroney M. J., Bookland R. G., Adang M. J. Molecular cloning and characterization of the insecticidal crystal protein gene of Bacillus thuringiensis var. tenebrionis. Proc Natl Acad Sci U S A. 1987 Oct;84(20):7036–7040. doi: 10.1073/pnas.84.20.7036. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Sick A., Gaertner F., Wong A. Nucleotide sequence of a coleopteran-active toxin gene from a new isolate of Bacillus thuringiensis subsp. tolworthi. Nucleic Acids Res. 1990 Mar 11;18(5):1305–1305. doi: 10.1093/nar/18.5.1305. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Thorne L., Garduno F., Thompson T., Decker D., Zounes M., Wild M., Walfield A. M., Pollock T. J. Structural similarity between the lepidoptera- and diptera-specific insecticidal endotoxin genes of Bacillus thuringiensis subsp. "kurstaki" and "israelensis". J Bacteriol. 1986 Jun;166(3):801–811. doi: 10.1128/jb.166.3.801-811.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Travers R. S., Martin P. A., Reichelderfer C. F. Selective Process for Efficient Isolation of Soil Bacillus spp. Appl Environ Microbiol. 1987 Jun;53(6):1263–1266. doi: 10.1128/aem.53.6.1263-1266.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Visser B., Munsterman E., Stoker A., Dirkse W. G. A novel Bacillus thuringiensis gene encoding a Spodoptera exigua-specific crystal protein. J Bacteriol. 1990 Dec;172(12):6783–6788. doi: 10.1128/jb.172.12.6783-6788.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Waalwijk C., Dullemans A. M., van Workum M. E., Visser B. Molecular cloning and the nucleotide sequence of the Mr 28 000 crystal protein gene of Bacillus thuringiensis subsp. israelensis. Nucleic Acids Res. 1985 Nov 25;13(22):8207–8217. doi: 10.1093/nar/13.22.8207. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Ward E. S., Ellar D. J. Nucleotide sequence of a Bacillus thuringiensis var. israelensis gene encoding a 130 kDa delta-endotoxin. Nucleic Acids Res. 1987 Sep 11;15(17):7195–7195. doi: 10.1093/nar/15.17.7195. [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. Yanisch-Perron C., Vieira J., Messing J. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene. 1985;33(1):103–119. doi: 10.1016/0378-1119(85)90120-9. [DOI] [PubMed] [Google Scholar]

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