Functional Characterizations of Residues Arg-158 and Tyr-170 of the Mosquito-larvicidal Bacillus Thuringiensis Cry4Ba

Overview

Journal BMB Rep

Specialties Biochemistry
Molecular Biology

Date 2013 Nov 30

PMID 24286331

Citations 1

Authors

Somphob Leetachewa

Saengduen Moonsom

Urai Chaisri

Narumol Khomkhum

Nonglak Yoonim

Ping Wang

Chanan Angsuthanasombat

Affiliations

Soon will be listed here.

Abstract

The insecticidal activity of Bacillus thuringiensis (Bt) Cry toxins involves toxin stabilization, oligomerization, passage across the peritrophic membrane (PM), binding to midgut receptors and pore-formation. The residues Arg-158 and Tyr-170 have been shown to be crucial for the toxicity of Bt Cry4Ba. We characterized the biological function of these residues. In mosquito larvae, the mutants R158A/E/Q (R158) could hardly penetrate the PM due to a significantly reduced ability to alter PM permeability; the mutant Y170A, however, could pass through the PM, but degraded in the space between the PM and the midgut epithelium. Further characterization by oligomerization demonstrated that Arg-158 mutants failed to form correctly sized high-molecular weight oligomers. This is the first report that Arg-158 plays a role in the formation of Cry4Ba oligomers, which are essential for toxin passage across the PM. Tyr-170, meanwhile, is involved in toxin stabilization in the toxic mechanism of Cry4Ba in mosquito larvae.

Citing Articles

Enhancement of insect susceptibility and larvicidal efficacy of Cry4Ba toxin by calcofluor.

Leetachewa S, Khomkhum N, Sakdee S, Wang P, Moonsom S Parasit Vectors. 2018; 11(1):515.

PMID: 30236155 PMC: 6148999. DOI: 10.1186/s13071-018-3110-3.

References

Likitvivatanavong S, Aimanova K, Gill S . Loop residues of the receptor binding domain of Bacillus thuringiensis Cry11Ba toxin are important for mosquitocidal activity. FEBS Lett. 2009; 583(12):2021-30. PMC: 2720071. DOI: 10.1016/j.febslet.2009.05.020. View

Bravo A, Gill S, Soberon M . Mode of action of Bacillus thuringiensis Cry and Cyt toxins and their potential for insect control. Toxicon. 2007; 49(4):423-35. PMC: 1857359. DOI: 10.1016/j.toxicon.2006.11.022. View

Jurat-Fuentes J, Adang M . Importance of Cry1 delta-endotoxin domain II loops for binding specificity in Heliothis virescens (L.). Appl Environ Microbiol. 2001; 67(1):323-9. PMC: 92576. DOI: 10.1128/AEM.67.1.323-329.2001. View

Halstead S . Dengue virus-mosquito interactions. Annu Rev Entomol. 2007; 53:273-91. DOI: 10.1146/annurev.ento.53.103106.093326. View

Rausell C, Munoz-Garay C, Miranda-CassoLuengo R, Gomez I, Rudino-Pinera E, Soberon M . 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. 2004; 43(1):166-74. DOI: 10.1021/bi035527d. View

Chayaratanasin P, Moonsom S, Sakdee S, Chaisri U, Katzenmeier G, Angsuthanasombat C . High level of soluble expression in Escherichia coli and characterisation of the cloned Bacillus thuringiensis Cry4Ba domain III fragment. J Biochem Mol Biol. 2007; 40(1):58-64. DOI: 10.5483/bmbrep.2007.40.1.058. View

Lee M, You T, Curtiss A, Dean D . 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. 1996; 229(1):139-46. DOI: 10.1006/bbrc.1996.1770. View

Lehane M . Peritrophic matrix structure and function. Annu Rev Entomol. 1997; 42:525-50. DOI: 10.1146/annurev.ento.42.1.525. View

EDWARDS . Permeability and disruption of the peritrophic matrix and caecal membrane from Aedes aegypti and Anopheles gambiae mosquito larvae. J Insect Physiol. 2000; 46(9):1313-1320. DOI: 10.1016/S0022-1910(00)00053-6. View

10.

Angsuthanasombat C, Uawithya P, Leetachewa S, Pornwiroon W, Ounjai P, Kerdcharoen T . Bacillus thuringiensis Cry4A and Cry4B mosquito-larvicidal proteins: homology-based 3D model and implications for toxin activity. J Biochem Mol Biol. 2004; 37(3):304-13. DOI: 10.5483/bmbrep.2004.37.3.304. View

11.

Vachon V, Prefontaine G, Rang C, Coux F, Juteau M, Schwartz J . Helix 4 mutants of the Bacillus thuringiensis insecticidal toxin Cry1Aa display altered pore-forming abilities. Appl Environ Microbiol. 2004; 70(10):6123-30. PMC: 522081. DOI: 10.1128/AEM.70.10.6123-6130.2004. View

12.

Leetachewa S, Katzenmeier G, Angsuthanasombat C . Novel preparation and characterization of the alpha4-loop-alpha5 membrane-perturbing peptide from the Bacillus thuringiensis Cry4Ba delta-endotoxin. J Biochem Mol Biol. 2006; 39(3):270-7. DOI: 10.5483/bmbrep.2006.39.3.270. View

13.

Gerber D, Shai Y . Insertion and organization within membranes of the delta-endotoxin pore-forming domain, helix 4-loop-helix 5, and inhibition of its activity by a mutant helix 4 peptide. J Biol Chem. 2000; 275(31):23602-7. DOI: 10.1074/jbc.M002596200. View

14.

Angsuthanasombat C, Crickmore N, Ellar D . Cytotoxicity of a cloned Bacillus thuringiensis subsp. israelensis CryIVB toxin to an Aedes aegypti cell line. FEMS Microbiol Lett. 1991; 67(3):273-6. DOI: 10.1016/0378-1097(91)90488-v. View

15.

Moskalyk L, Oo M, Jacobs-Lorena M . Peritrophic matrix proteins of Anopheles gambiae and Aedes aegypti. Insect Mol Biol. 1996; 5(4):261-8. DOI: 10.1111/j.1365-2583.1996.tb00100.x. View

16.

Kanintronkul Y, Sramala I, Katzenmeier G, Panyim S, Angsuthanasombat C . Specific mutations within the alpha4-alpha5 loop of the Bacillus thuringiensis Cry4B toxin reveal a crucial role for Asn-166 and Tyr-170. Mol Biotechnol. 2003; 24(1):11-20. DOI: 10.1385/MB:24:1:11. View

17.

Khaokhiew T, Angsuthanasombat C, Promptmas C . Correlative effect on the toxicity of three surface-exposed loops in the receptor-binding domain of the Bacillus thuringiensis Cry4Ba toxin. FEMS Microbiol Lett. 2009; 300(1):139-45. DOI: 10.1111/j.1574-6968.2009.01774.x. View

18.

Borovsky D, Meola S . Biochemical and cytoimmunological evidence for the control of Aedes aegypti larval trypsin with Aea-TMOF. Arch Insect Biochem Physiol. 2004; 55(3):124-39. DOI: 10.1002/arch.10132. View

19.

Ito T, Bando H, Asano S . Activation process of the mosquitocidal delta-endotoxin Cry39A produced by Bacillus thuringiensis subsp. aizawai BUN1-14 and binding property to Anopheles stephensi BBMV. J Invertebr Pathol. 2006; 93(1):29-35. DOI: 10.1016/j.jip.2006.05.007. View

20.

Boonserm P, Davis P, Ellar D, Li J . Crystal structure of the mosquito-larvicidal toxin Cry4Ba and its biological implications. J Mol Biol. 2005; 348(2):363-82. DOI: 10.1016/j.jmb.2005.02.013. View