Tags

Type your tag names separated by a space and hit enter

Mapping of the entomocidal fragment of Spodoptera-specific Bacillus thuringiensis toxin CryIC.
Mol Gen Genet. 1996 Nov 27; 253(1-2):11-9.MG

Abstract

Insecticidal CryI protoxins of Bacillus thuringiensis are activated by proteolysis in the midgut of insects. A conservation of proteolytic cleavage sites in the CryI proteins facilitates the expression of active toxins in transgenic plants to obtain protection from various insects. However, the engineering of CryIC toxins has, thus far, failed to yield applicable resistance to armyworms of Spodoptera species representing common insect pests worldwide. To improve the production of recombinant CryIC toxins, we established a CryIC consensus sequence by comparative analysis of three cryIC genes and tested the stability and protease sensitivity of truncated CryIC toxins in Escherichia coli and in vitro. In contrast to previous data, the boundaries of trypsin-resistant CryIC core toxin were mapped to amino acid residues I28 and R627. Proteolysis of the truncated CryIC proteins showed that Spodoptera midgut proteases may further shorten the C-terminus of CryIC toxin to residue A615. However, C-terminal truncation of CryIC to residue L614, and a mutation causing amino acid replacement I610T, abolished the insecticidal activity of CryIC toxin to S. littoralis larvae, as well as its resistance to trypsin and Spodoptera midgut proteases. Because no CryIC toxin carrying a proteolytically processed N-terminus could be stably expressed in bacteria, our data indicate that, in contrast to other CryI proteins, an entomocidal fragment located between amino acid positions 1 and 627 is required for stable production of recombinant CryIC toxins.

Authors+Show Affiliations

Strizhov N
Max-Planck Institut für Züchtungsforschung, Köln, Germany.
Keller M
No affiliation info available
Koncz-Kálmán Z
No affiliation info available
Regev A
No affiliation info available
Sneh B
No affiliation info available
Schell J
No affiliation info available
Koncz C
No affiliation info available
Zilberstein A
No affiliation info available
Konez-Kálmán Z
No affiliation info available

MeSH

Amino Acid SequenceAnimalsBacillus thuringiensisBacillus thuringiensis ToxinsBacterial ProteinsBacterial ToxinsCloning, MolecularConsensus SequenceEndopeptidasesEndotoxinsGenes, BacterialHemolysin ProteinsInsecticidesMolecular Sequence DataMutationProtein Processing, Post-TranslationalRecombinant ProteinsSpodopteraTrypsin

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

9003281

Citation

Strizhov, N, et al. "Mapping of the Entomocidal Fragment of Spodoptera-specific Bacillus Thuringiensis Toxin CryIC." Molecular & General Genetics : MGG, vol. 253, no. 1-2, 1996, pp. 11-9.
Strizhov N, Keller M, Koncz-Kálmán Z, et al. Mapping of the entomocidal fragment of Spodoptera-specific Bacillus thuringiensis toxin CryIC. Mol Gen Genet. 1996;253(1-2):11-9.
Strizhov, N., Keller, M., Koncz-Kálmán, Z., Regev, A., Sneh, B., Schell, J., Koncz, C., Zilberstein, A., & Konez-Kálmán, Z. (1996). Mapping of the entomocidal fragment of Spodoptera-specific Bacillus thuringiensis toxin CryIC. Molecular & General Genetics : MGG, 253(1-2), 11-9.
Strizhov N, et al. Mapping of the Entomocidal Fragment of Spodoptera-specific Bacillus Thuringiensis Toxin CryIC. Mol Gen Genet. 1996 Nov 27;253(1-2):11-9. PubMed PMID: 9003281.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Mapping of the entomocidal fragment of Spodoptera-specific Bacillus thuringiensis toxin CryIC. AU - Strizhov,N, AU - Keller,M, AU - Koncz-Kálmán,Z, AU - Regev,A, AU - Sneh,B, AU - Schell,J, AU - Koncz,C, AU - Zilberstein,A, AU - Konez-Kálmán,Z, PY - 1996/11/27/pubmed PY - 2000/3/23/medline PY - 1996/11/27/entrez SP - 11 EP - 9 JF - Molecular & general genetics : MGG JO - Mol Gen Genet VL - 253 IS - 1-2 N2 - Insecticidal CryI protoxins of Bacillus thuringiensis are activated by proteolysis in the midgut of insects. A conservation of proteolytic cleavage sites in the CryI proteins facilitates the expression of active toxins in transgenic plants to obtain protection from various insects. However, the engineering of CryIC toxins has, thus far, failed to yield applicable resistance to armyworms of Spodoptera species representing common insect pests worldwide. To improve the production of recombinant CryIC toxins, we established a CryIC consensus sequence by comparative analysis of three cryIC genes and tested the stability and protease sensitivity of truncated CryIC toxins in Escherichia coli and in vitro. In contrast to previous data, the boundaries of trypsin-resistant CryIC core toxin were mapped to amino acid residues I28 and R627. Proteolysis of the truncated CryIC proteins showed that Spodoptera midgut proteases may further shorten the C-terminus of CryIC toxin to residue A615. However, C-terminal truncation of CryIC to residue L614, and a mutation causing amino acid replacement I610T, abolished the insecticidal activity of CryIC toxin to S. littoralis larvae, as well as its resistance to trypsin and Spodoptera midgut proteases. Because no CryIC toxin carrying a proteolytically processed N-terminus could be stably expressed in bacteria, our data indicate that, in contrast to other CryI proteins, an entomocidal fragment located between amino acid positions 1 and 627 is required for stable production of recombinant CryIC toxins. SN - 0026-8925 UR - https://www.unboundmedicine.com/medline/citation/9003281/Mapping_of_the_entomocidal_fragment_of_Spodoptera_specific_Bacillus_thuringiensis_toxin_CryIC_ L2 - https://doi.org/10.1007/s004380050290 DB - PRIME DP - Unbound Medicine ER -