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. 1997 Jul;179(13):4336–4341. doi: 10.1128/jb.179.13.4336-4341.1997

Cloning and analysis of the first cry gene from Bacillus popilliae.

J Zhang 1, T C Hodgman 1, L Krieger 1, W Schnetter 1, H U Schairer 1
PMCID: PMC179258  PMID: 9209052

Abstract

An 80-kDa parasporal crystal protein was detected in protein extracts of sporangia of Bacillus popilliae isolated from a diseased larva of the common cockchafer (Melolontha melolontha L.). Amino acid analysis of tryptic peptides revealed significant homology to the Cry2Aa endotoxins of Bacillus thuringiensis. The gene cryBP1 (cry18Aa1), which codes for the parasporal crystal protein, was found in a putative cry operon on the bacterial chromosome, which contains at least one further (smaller) open reading frame, orf1. The 706-amino-acid-long CryBP1 (Cry18Aa1) protein has a predicted molecular mass of 79 kDa and shows about 40% sequence identity to the Cry2 polypeptides of B. thuringiensis. In the light of published observations which suggest that the parasporal crystal proteins of B. popilliae are slightly toxic to their grub hosts, we propose the following survival strategy of B. popilliae. As an obligate pathogen of grubs, B. popilliae germinates in the gut of a grub and the parasporal crystal proteins are released and activated. The activated protein does not cause colloid osmotic lysis but instead damages the gut wall somehow to allow the vegetative cells to enter the hemolymph more easily. By becoming a parasite, B. popilliae can continue to proliferate efficiently while the living grub provides a food supply. This process is in contrast to that of B. thuringiensis, which rapidly kills the insect and is then limited to growth on the larval carcass.

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Selected References

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  1. Adams L. F., Visick J. E., Whiteley H. R. A 20-kilodalton protein is required for efficient production of the Bacillus thuringiensis subsp. israelensis 27-kilodalton crystal protein in Escherichia coli. J Bacteriol. 1989 Jan;171(1):521–530. doi: 10.1128/jb.171.1.521-530.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Ahmad W., Ellar D. J. Directed mutagenesis of selected regions of a Bacillus thuringiensis entomocidal protein. FEMS Microbiol Lett. 1990 Mar 1;56(1-2):97–104. doi: 10.1016/0378-1097(90)90132-a. [DOI] [PubMed] [Google Scholar]
  3. Altschul S. F., Lipman D. J. Protein database searches for multiple alignments. Proc Natl Acad Sci U S A. 1990 Jul;87(14):5509–5513. doi: 10.1073/pnas.87.14.5509. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Barloy F., Delécluse A., Nicolas L., Lecadet M. M. Cloning and expression of the first anaerobic toxin gene from Clostridium bifermentans subsp. malaysia, encoding a new mosquitocidal protein with homologies to Bacillus thuringiensis delta-endotoxins. J Bacteriol. 1996 Jun;178(11):3099–3105. doi: 10.1128/jb.178.11.3099-3105.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Brown K. L., Whiteley H. R. Isolation of a Bacillus thuringiensis RNA polymerase capable of transcribing crystal protein genes. Proc Natl Acad Sci U S A. 1988 Jun;85(12):4166–4170. doi: 10.1073/pnas.85.12.4166. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Dankocsik C., Donovan W. P., Jany C. S. Activation of a cryptic crystal protein gene of Bacillus thuringiensis subspecies kurstaki by gene fusion and determination of the crystal protein insecticidal specificity. Mol Microbiol. 1990 Dec;4(12):2087–2094. doi: 10.1111/j.1365-2958.1990.tb00569.x. [DOI] [PubMed] [Google Scholar]
  7. Dervyn E., Poncet S., Klier A., Rapoport G. Transcriptional regulation of the cryIVD gene operon from Bacillus thuringiensis subsp. israelensis. J Bacteriol. 1995 May;177(9):2283–2291. doi: 10.1128/jb.177.9.2283-2291.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. 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]
  9. 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]
  10. Ge A. Z., Shivarova N. I., Dean D. H. Location of the Bombyx mori specificity domain on a Bacillus thuringiensis delta-endotoxin protein. Proc Natl Acad Sci U S A. 1989 Jun;86(11):4037–4041. doi: 10.1073/pnas.86.11.4037. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Grochulski P., Masson L., Borisova S., Pusztai-Carey M., Schwartz J. L., Brousseau R., Cygler M. Bacillus thuringiensis CryIA(a) insecticidal toxin: crystal structure and channel formation. J Mol Biol. 1995 Dec 1;254(3):447–464. doi: 10.1006/jmbi.1995.0630. [DOI] [PubMed] [Google Scholar]
  12. Göbel U., Maas R., Clad A. Quantitative electroelution of oligonucleotides and large DNA fragments from gels and purification by electrodialysis. J Biochem Biophys Methods. 1987 Aug;14(5):245–260. doi: 10.1016/0165-022x(87)90050-9. [DOI] [PubMed] [Google Scholar]
  13. 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]
  14. Hofmann C., Lüthy P., Hütter R., Pliska V. Binding of the delta endotoxin from Bacillus thuringiensis to brush-border membrane vesicles of the cabbage butterfly (Pieris brassicae). Eur J Biochem. 1988 Apr 5;173(1):85–91. doi: 10.1111/j.1432-1033.1988.tb13970.x. [DOI] [PubMed] [Google Scholar]
  15. 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]
  16. 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]
  17. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  18. Li J. D., Carroll J., Ellar D. J. Crystal structure of insecticidal delta-endotoxin from Bacillus thuringiensis at 2.5 A resolution. Nature. 1991 Oct 31;353(6347):815–821. doi: 10.1038/353815a0. [DOI] [PubMed] [Google Scholar]
  19. Macdonald R., Kalmakoff J. Comparison of Pulsed-Field Gel Electrophoresis DNA Fingerprints of Field Isolates of the Entomopathogen Bacillus popilliae. Appl Environ Microbiol. 1995 Jun;61(6):2446–2449. doi: 10.1128/aem.61.6.2446-2449.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Meade H. M., Long S. R., Ruvkun G. B., Brown S. E., Ausubel F. M. Physical and genetic characterization of symbiotic and auxotrophic mutants of Rhizobium meliloti induced by transposon Tn5 mutagenesis. J Bacteriol. 1982 Jan;149(1):114–122. doi: 10.1128/jb.149.1.114-122.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Valyasevi R., Kyle M. M., Christie P. J., Steinkraus K. H. Plasmids in Bacillus popilliae. J Invertebr Pathol. 1990 Sep;56(2):286–288. doi: 10.1016/0022-2011(90)90113-k. [DOI] [PubMed] [Google Scholar]
  22. Van Rie J., Jansens S., Höfte H., Degheele D., Van Mellaert H. Receptors on the brush border membrane of the insect midgut as determinants of the specificity of Bacillus thuringiensis delta-endotoxins. Appl Environ Microbiol. 1990 May;56(5):1378–1385. doi: 10.1128/aem.56.5.1378-1385.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Walters F. S., Slatin S. L., Kulesza C. A., English L. H. Ion channel activity of N-terminal fragments from CryIA(c) delta-endotoxin. Biochem Biophys Res Commun. 1993 Oct 29;196(2):921–926. doi: 10.1006/bbrc.1993.2337. [DOI] [PubMed] [Google Scholar]
  24. Weiner B. A. Isolation and partial characterization of the parasporal body of Bacillus popilliae. Can J Microbiol. 1978 Dec;24(12):1557–1561. doi: 10.1139/m78-249. [DOI] [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. Wu D., Aronson A. I. Localized mutagenesis defines regions of the Bacillus thuringiensis delta-endotoxin involved in toxicity and specificity. J Biol Chem. 1992 Feb 5;267(4):2311–2317. [PubMed] [Google Scholar]
  27. Wu D., Cao X. L., Bai Y. Y., Aronson A. I. Sequence of an operon containing a novel delta-endotoxin gene from Bacillus thuringiensis. FEMS Microbiol Lett. 1991 Jun 1;65(1):31–35. doi: 10.1016/0378-1097(91)90466-n. [DOI] [PubMed] [Google Scholar]
  28. de Maagd R. A., Kwa M. S., van der Klei H., Yamamoto T., Schipper B., Vlak J. M., Stiekema W. J., Bosch D. Domain III substitution in Bacillus thuringiensis delta-endotoxin CryIA(b) results in superior toxicity for Spodoptera exigua and altered membrane protein recognition. Appl Environ Microbiol. 1996 May;62(5):1537–1543. doi: 10.1128/aem.62.5.1537-1543.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]

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