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| The Minimal Active Fragment of the Cry1Ai Toxin is Located Between 36I and 605I |
| ZHOU Zi-shan, LIN Hui-yan, LI Ying, SHU Chang-long, SONG Fu-ping , ZHANG Jie |
| State Key Laboratory for Biology of Plant Diseases and Insect Pests/Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, P.R.China |
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摘要 The novel cry1Ai gene that cloned from Bacillus thuringiensis strain SC6H8 encoded a protein exhibiting strong toxicity against Plutella xylostella and Chilo suppressalis in our previous study. Using the available information for the active fragments of other Cry toxins, eight truncated fragments were constructed to identify the minimal active fragment of Cry1Ai. All truncated fragments were expressed in Escherichia coli strain BL21 (DE3), and the insecticidal activity against 2nd- instar P. xylostella larvae was assessed using full-length Cry1Ai as a positive control. The results indicate that the minimal active fragment of the Cry1Ai toxin against P. xylostella is located between amino acid residues 36I and 605I, which is smaller than the regions previously reported for Cry1A. The first two amino acids (34T and 35P) on helix α-1 and whole helix α-2 of domain I and sheet β-32 of domain III are necessary for Cry1Ai toxin to keep its toxicity against P. xylostella.
Abstract The novel cry1Ai gene that cloned from Bacillus thuringiensis strain SC6H8 encoded a protein exhibiting strong toxicity against Plutella xylostella and Chilo suppressalis in our previous study. Using the available information for the active fragments of other Cry toxins, eight truncated fragments were constructed to identify the minimal active fragment of Cry1Ai. All truncated fragments were expressed in Escherichia coli strain BL21 (DE3), and the insecticidal activity against 2nd- instar P. xylostella larvae was assessed using full-length Cry1Ai as a positive control. The results indicate that the minimal active fragment of the Cry1Ai toxin against P. xylostella is located between amino acid residues 36I and 605I, which is smaller than the regions previously reported for Cry1A. The first two amino acids (34T and 35P) on helix α-1 and whole helix α-2 of domain I and sheet β-32 of domain III are necessary for Cry1Ai toxin to keep its toxicity against P. xylostella.
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Received: 20 March 2013
Accepted:
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| Fund: This study was supported by the National Natural ScienceFoundation of China (31272115), the National 973 Program of China (2009CB118902). We thank Ms. Liang Yingping and Liu Shuqin from Institution of Plant Protection, Chinese Academy of Agricultural Sciences, for their assistance with insect rearing and the bioassays. |
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Corresponding Authors:
ZHANG Jie, Tel: +86-10-62815921, Fax: +86-10-62812642, E-mail: jzhang@ippcaas.cn
E-mail: jzhang@ippcaas.cn
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Cite this article:
ZHOU Zi-shan, LIN Hui-yan, LI Ying, SHU Chang-long, SONG Fu-ping , ZHANG Jie.
2014.
The Minimal Active Fragment of the Cry1Ai Toxin is Located Between 36I and 605I. Journal of Integrative Agriculture, 13(5): 1036-1042.
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| Arnold S, Curtiss A, Dean D H, Alzate O. 2001. The role of a proline-induced broken-helix motif in α-helix 2 of Bacillus thuringiensis δ-endotoxins. FEBS Letters, 490, 70-74 Bietlot H P, Carey P R, Pozsgay M, Kaplan H. 1989. Isolation of carboxyl-terminal peptides from proteins by diagonal electrophoresis: Application to the entomocidal toxin from Bacillus thuringiensis. Analytical Biochemistry, 181, 212-215 Bravo A, Gómez I, Conde J, Munoz-Garay C, Sánchez J, Miranda R, Zhuang M, Gill S S, Soberón M. 2004. Oligomerization triggers binding of a Bacillus thuringiensis Cry1Ab pore-forming toxin to aminopeptidase N receptor leading to insertion into membrane microdomains. Biochimica et Biophysica Acta-Biomembranes, 1667, 38-46 Bravo A, Gómez I, Porta H, García-Gómez B I, Rodriguez- Almazan C, Pardo L, Soberón M. 2012. Evolution of Bacillus thuringiensis Cry toxins insecticidal activity. Microbial Biotechnology, 6, 17-26 Bravo A, Likitvivatanavong S, Gill S S, Soberón M. 2011. Bacillus thuringiensis: A story of a successful bioinsecticide. Insect Biochemistry and Molecular Biology, 41, 423-431 Chen M, Zhao J Z, Collins H L, Earle E D, Cao J, Shelton A M. 2008. A critical assessment of the effects of Bt transgenic plants on parasitoids. PLoS One, 3, e2284. Griffitts J S, Huffman D L, Whitacre J L, Barrows B D, Marroquin L D, Müller R, Brown J R, Hennet T, Esko J D, Aroian R V. 2003. Resistance to a bacterial toxin is mediated by removal of a conserved glycosylation pathway required for toxin-host interactions. The Journal of Biological Chemistry, 278, 5594-45602 Jiménez-Juárez N, Munoz-Garay C, Gómez I, SaabRincon G, Damian-Alamazo J Y, Gill S S, Soberón M, Bravo A. 2007. Bacillus thuringiensis Cry1Ab mutants affecting oligomer formation are non toxic to Manduca sexta larvae. Journal of Biological Chemistry, 282, 21222- 21229. Lee M K, Miles P, Chen J S. 2006. Brush border membrane binding properties of Bacillus thuringiensis Vip3A toxin to Heliothis virescens and Helicoverpa zea midguts. Biochemical and Biophysical Research Communications, 339, 1043-1047 Mendelson M, Kough J, Vaituzis Z, Matthews K. 2003. Are Bt crops safe? Nature Biotechnology, 21, 1003-1009 Nagamatsu Y, Itai Y, Hatanaka C, Funatsu G, Hayashi K. 1984. A toxic fragment from the entomocidal crystal protein of Bacillus thuringiensis. Agricultural and Biological Chemistry, 48, 611-619 Ohsawa M, Tanaka M, Moriyama K, Shimazu M, Asano S, Miyamoto K, Haginoya K, Mitsui T, Kouya T, Taniguchi M, Horia H. 2012. A 50-kilodalton Cry2A peptide is lethal to Bombyx mori and Lymantria dispar. Applied and Environmental Microbiology, 78, 4755- 4757. Pacheco S, Gómez I, Arenas I, Saab-Rincon G, Rodriguez- Almazan C, Gill S S, Bravo A, Soberón M. 2009. Domain II loop 3 of Bacillus thuringiensis Cry1Ab toxin is involved in a “ping-pong” binding mechanism with Manduca sexta aminopetidase-N and cadherin receptors. Journal of Biological Chemistry, 284, 32750-32757 Pardo-López L, Soberón M, Bravo A. 2013. Bacillus thuringiensis insecticidal three-domain Cry toxins: mode of action, insect resistance and consequences for crop protection. FEMS Microbiology Reviews, 37, 3-22 Pigott C R, Ellar D J. 2007. Role of receptors in Bacillus thuringiensis crystal toxin activity. Microbiology and Molecular Biology Reviews, 71, 255-281 Qaim M, Zilberman D. 2003. Yield effects of genetically modified crops in developing countries. Science, 299, 900-902 Rodríguez-Almazán C, Zavala L E, Munoz-Garay C, Jiménez-Juárez N, Pacheco S, Masson L, Soberón M, Bravo A. 2009. Dominant negative mutants of Bacillus thuringiensis Cry1Ab toxin function as anti-toxins: Demonstration of the role of oligomerization in toxicity. The Public Library of Science One, 4, e5545.Sambrook J, Russell D W. 2002. Molecular Cloning: A Laboratory Manual. 3rd ed. Science Press, Beijing. (in Chinese) Schnepf E, Crickmore N, Van R J, Lereclus D, Baum J, Feitelson J, Zeigler D R, Dean D H. 1998. Bacillus thuringiensis and its pesticidal crystal proteins. Microbiology and Molecular Biology Reviews, 62, 775- 806. Schnepf H E, Whiteley H R. 1981. Cloning and expression of the Bacillus thuringiensis crystal protein gene in Escherichia coli. Proceedings of the National Academy of Sciences of the United States of America, 78, 2893- 2897. Shu C L, Song F P, Zhang J, Huang D F, He K L, Liang G M. 2013. The Application of Bacillus Thuringiensis Cry1ai Gene, and Modified Mcry1ai Gene. China Patent, No. ZL200910241558.8. Soberón M, Pardo-López L, López I, Gómez I, Tabashnik B, Bravo A. 2007. Engineering modified Bt toxins to counter insect resistance. Science, 318, 1640-1642 Strizhov N, Keller M, Mathur J, Koncz-Kálmán Z, Bosch D, Prudovsky E, Schell J, Sneh B, Koncz C, Zilberstein A. 1996. A synthetic cryIC gene, encoding a Bacillus thuringiensis δ-endotoxin, confers Spodoptera resistance in alfalfa and tobacco. Proceedings of the National Academy of Sciences of the United States of America, 93, 15012-15017 Tabashnik B E, Huang F, Ghimire M N, Leonard B R, Siegfried B D, Rangasamy M, Soberón M. 2011. Efficacy of genetically modified Bt toxins against insects with different genetic mechanisms of resistance. Nature Biotechnology, 29, 1128-1131 Tigue N J, Jacoby J, Ellar D J. 2001. The α-Helix 4 residue, Asn135, is involved in the oligomerization of Cry1Ac1 and Cry1Ab5 Bacillus thuringiensistoxins. Applied and Environmental Microbiology, 67, 5715-5720 Vaeck M, Reynaerts A, Hofte H, Jansens S, de Beuckeleer M, Dean C, Zabeau M, van Montagu M, Leemans J. 1987. Transgenic plants protected from insect attack. Nature, 328, 33-37 Wang G J, Zhang J, Wu J, Song F P, Huang D F. 2005. Toxicity analysis of truncated insecticidal crystal protein Cry1Ba3 from Bacillus thuringiensis. Scientia Agricultura Sinica, 38, 1585-1590 (in Chinese) Wu Y E, Zhang J, Cao J P, Huang D F, Song F P. 2003. Study on truncated CrylIel protein from Bacillus thuringiensis. Plant Protection, 29, l5-l8. (in Chinese) Xue J, Zhou Z S, Song F P, Shu C L, Huang D F, Zhang J. 2011. Identification of the minimal active fragment of the Cry1Ah toxin. Biotechnology Letters, 33, 531-537 Zaidi M, Ye G, Yao H, You T, Loit E, Dean D, Riazuddin S, Altosaar I. 2009. Transgenic rice plants expressing a modified gene are resistant to Spodoptera litura and Chilo suppressalis. Molecular Biotechnology, 43, 232- 242. Zhang J T, Shu C L, Song F P, Liu N, Zhang J, Huang D F. 2009. Cloning, expression and insecticidal activity of cry2A gene from Bacillus thuringiensis. Biotechnology Bulletin, 10, 145-150 (in Chinese) |
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