Functional characterizations of residues Arg-158 and Tyr-170 of the mosquito-larvicidal Bacillus thuringiensis Cry4Ba
1
조회
0
다운로드
0
인용
0
참고문헌
키워드
초록
The insecticidal activity of Bacillus thuringiensis (Bt) Cry toxinsinvolves toxin stabilization, oligomerization, passage acrossthe peritrophic membrane (PM), binding to midgut receptorsand pore-formation. The residues Arg-158 and Tyr-170 havebeen shown to be crucial for the toxicity of Bt Cry4Ba. Wecharacterized the biological function of these residues. Inmosquito larvae, the mutants R158A/E/Q (R158) could hardlypenetrate the PM due to a significantly reduced ability to alterPM permeability; the mutant Y170A, however, could passthrough the PM, but degraded in the space between the PMand the midgut epithelium. Further characterization byoligomerization demonstrated that Arg-158 mutants failed toform correctly sized high-molecular weight oligomers. This isthe first report that Arg-158 plays a role in the formation ofCry4Ba oligomers, which are essential for toxin passage acrossthe PM. Tyr-170, meanwhile, is involved in toxin stabilization in the toxic mechanism of Cry4Ba in mosquito larvae.
Abstract
The insecticidal activity of Bacillus thuringiensis (Bt) Cry toxinsinvolves toxin stabilization, oligomerization, passage acrossthe peritrophic membrane (PM), binding to midgut receptorsand pore-formation. The residues Arg-158 and Tyr-170 havebeen shown to be crucial for the toxicity of Bt Cry4Ba. Wecharacterized the biological function of these residues. Inmosquito larvae, the mutants R158A/E/Q (R158) could hardlypenetrate the PM due to a significantly reduced ability to alterPM permeability; the mutant Y170A, however, could passthrough the PM, but degraded in the space between the PMand the midgut epithelium. Further characterization byoligomerization demonstrated that Arg-158 mutants failed toform correctly sized high-molecular weight oligomers. This isthe first report that Arg-158 plays a role in the formation ofCry4Ba oligomers, which are essential for toxin passage acrossthe PM. Tyr-170, meanwhile, is involved in toxin stabilization in the toxic mechanism of Cry4Ba in mosquito larvae.
참고문헌 (27)
- Bravo, A., "Mode of action of Bacillus thuringiensis Cry and Cyt toxins and their potential for insect control", Toxicon, 49, pp.423~435 (2007)
- Ito, T., "Activation process of the mosquitocidal δ-endotoxin Cry39A produced by Bacillus thuringiensis subsp. aizawai BUN1-14 and binding property to Anopheles stephensi BBMV", J. Invertebr. Pathol, 93, pp.29~35 (2006)
- Angsuthanasombat, C., "Cytotoxicity of a cloned Bacillus thuringiensis subsp. israelensis CryIVB toxin to an Aedes aegypti cell line", FEMS Microbiol. Lett, 67, pp.273~276 (1991)
- Halstead, S. B., "Dengue virus-mosquito interactions", Annu. Rev. Entomol, 53, pp.273~291 (2008)
- Angsuthanasombat, C., "Bacillus thuringiensis Cry4A and Cry4B mosquito-larvicidal proteins: homology- based 3D model and implications for toxin activity", J. Biochem. Mol. Biol, 37, pp.304~313 (2004)
- Boonserm, P., "Crystal structure of the mosquito-larvicidal toxin Cry4Ba and its biological implications", J. Mol. Biol, 348, pp.363~382 (2005)
- Knowles, B. H., "Colloid-osmotic lysis is a general feature of the mechanism of action of Bacillus thuringiensis δ-endotoxins with different insect specificity", Biochimica et Biophysica Acta (BBA) - General Subjects, 924, pp.509~518 (1987)
- Gerber, D., "Insertion and organization within membranes of the δ-endotoxin pore-forming domain, helix 4-loop-helix 5, and inhibition of its activity by a mutant helix 4 peptide", J. Biol. Chem, 275, pp.23602~23607 (2000)
- Leetachewa, S., "Novel preparation and characterization of the α4-loop-α5 membrane-perturbing peptide from the Bacillus thuringiensis Cry4Ba δ-endotoxin", J. Biochem. Mol. Biol, 39, pp.270~277 (2006)
- Masson, L., "Helix 4 of the Bacillus thuringiensis Cry1Aa toxin lines the lumen of the ion channel", J. Biol. Chem, 274, pp.31996~32000 (1999)
- Vachon, V., "Helix 4 mutants of the Bacillus thuringiensis insecticidal toxin Cry1Aa display altered pore-forming abilities", Appl. Environ. Microbiol, 70, pp.6123~6130 (2004)
- Sramala, I., "Charged residues screening in helix 4 of the Bacillus thuringiensis Cry4B toxin reveals one critical residue for larvicidal activity", J. Biochem. Mol. Biol. Biophys, 5, pp.219~215 (2001)
- Sramala, I., "Institute of Molecular Biology and Genetics", pp.232~ (2002), Mahidol University, Nakhon Pathom
- Kanintronkul, Y., "Specific mutations within the α4-α5 loop of the Bacillus thuringiensis Cry4B toxin reveal a crucial role for Asn-166 and Tyr-170", Mol. Biotechnol, 24, pp.11~20 (2003)
- Lehane, M. J., "Peritrophic matrix structure and function", Annu. Rev. Entomol, 42, pp.525~550 (1997)
- Peters, W., "Peritrophic membranes" (1992), Springer
- Jurat-Fuentes, J. L., "Importance of Cry1 δ-endotoxin domain II loops for binding specificity in Heliothis virescens (L.)", Appl. Environ. Microbiol, 67, pp.323~329 (2001)
- Lee, M. K., "Involvement of two amino acid residues in the loop region of Bacillus thuringiensis Cry1Ab toxin in toxicity and binding to Lymantria dispar", Biochem. Biophys. Res. Commun, 229, pp.139~146 (1996)
- Khaokhiew, T., "Correlative effect on the toxicity of three surface- exposed loops in the receptor-binding domain of the Bacillus thuringiensis Cry4Ba toxin", FEMS Microbiol. Lett, 300, pp.139~145 (2009)
- Tuntitippawan, T., "Targeted mutagenesis of loop residues in the receptor-binding domain of the Bacillus thuringiensis Cry4Ba toxin affects larvicidal activity", FEMS Microbiol. Lett, 242, pp.325~332 (2005)
- Likitvivatanavong, S., "Loop residues of the receptor binding domain of Bacillus thuringiensis Cry11Ba toxin are important for mosquitocidal activity", FEBS Lett, 583, pp.2021~2030 (2009)
- Borovsky, D., "Biochemical and cytoimmunological evidence for the control of Aedes aegypti larval trypsin with Aea-TMOF", Arch. Insect Biochem. Physiol, 55, pp.124~139 (2004)
- Rausell, C., "Tryptophan spectroscopy studies and black lipid bilayer analysis indicate that the oligomeric structure of Cry1Ab toxin from Bacillus thuringiensis is the membrane- insertion intermediate", Biochemistry, 43, pp.166~174 (2004)
- Moskalyk, L. A., "Peritrophic matrix proteins of Anopheles gambiae and Aedes aegypti", Insect Mol. Biol, 5, pp.261~268 (1996)
- Edwards, M. J., "Permeability and disruption of the peritrophic matrix and caecal membrane from Aedes aegypti and Anopheles gambiae mosquito larvae", J. Insect. Physiol, 46, pp.1313~1320 (2000)
- Moonsom, S., "Molecular Genetics and Genetic Engineering, Faculty of Graduate Studies", pp.133~ (2004), Mahidol University, Nakhon Pathom
- Chayaratanasin, P., "High level of soluble expression in Escherichia coli and characterisation of the cloned Bacillus thuringiensis Cry4Ba domain III fragment", J. Biochem. Mol. Biol, 40, pp.58~64 (2007)