Composition of the Putative Prepore Complex of Bacillus thuringiensis Cry1Ab Toxin
Affiliation(s)
1 Department of Chemistry and Biochemistry, The Ohio State University, Columbus, OH, USA. 2 Aaron Diamond AIDS Research Center, Rockefeller University, New York, NY, USA.
1 Department of Chemistry and Biochemistry, The Ohio State University, Columbus, OH, USA. 2 Aaron Diamond AIDS Research Center, Rockefeller University, New York, NY, USA.
ABSTRACT
Prepore formation is hypothesized to be an obligate step in the insertion of Cry1Ab toxin into insect brush border membrane vesicles. We examined the architecture of the putative prepore when isolated using the published protocols [1] [2]. Our results demonstrate that the putative prepore form of Cry1Ab is a combination of receptor proteins attached to the toxin, when purified. The results also suggest that this prepore form as prepared by the methods published is different from other membrane-extracted oligomeric forms of Cry toxins and prepore of other toxins in general. While most other known prepores are composed of multimers of a single protein, the Cry1Ab prepore, as generated, is a protein-receptor complex oligomer and monomers of Cry toxins.
Prepore formation is hypothesized to be an obligate step in the insertion of Cry1Ab toxin into insect brush border membrane vesicles. We examined the architecture of the putative prepore when isolated using the published protocols [1] [2]. Our results demonstrate that the putative prepore form of Cry1Ab is a combination of receptor proteins attached to the toxin, when purified. The results also suggest that this prepore form as prepared by the methods published is different from other membrane-extracted oligomeric forms of Cry toxins and prepore of other toxins in general. While most other known prepores are composed of multimers of a single protein, the Cry1Ab prepore, as generated, is a protein-receptor complex oligomer and monomers of Cry toxins.
KEYWORDS
Cry1Ab Protein, Protein-Receptor Interactions, Brush Border Membrane Vesicles, Small Unilamellar Vesicles, Prepore, LC-MS/MS
Cry1Ab Protein, Protein-Receptor Interactions, Brush Border Membrane Vesicles, Small Unilamellar Vesicles, Prepore, LC-MS/MS
Cite this paper
Nair, M. and Dean, D. (2015) Composition of the Putative Prepore Complex of Bacillus thuringiensis Cry1Ab Toxin. Advances in Biological Chemistry, 5, 179-188. doi: 10.4236/abc.2015.54014.
Nair, M. and Dean, D. (2015) Composition of the Putative Prepore Complex of Bacillus thuringiensis Cry1Ab Toxin. Advances in Biological Chemistry, 5, 179-188. doi: 10.4236/abc.2015.54014.
References
[1] Gomez, I., Sanchez, J., Miranda, R., Bravo, A. and Soberon, M. (2002) Cadherin-Like Receptor Binding Facilitates Proteolytic Cleavage of Helix Alpha-1 in Domain I and Oligomer Pre-Pore Formation of Bacillus thuringiensis Cry1Ab Toxin. FEBS Letters, 513, 242-246.
http://dx.doi.org/10.1016/S0014-5793(02)02321-9 [2] Jimenez-Juarez, N., Munoz-Garay, C., Gomez, I., Saab-Rincon, G., Damian-Almazo, J.Y., Gill, S.S., Soberon, M. and Bravo, A. (2007) Bacillus thuringiensis Cry1Ab Mutants Affecting Oligomer Formation Are Non-Toxic to Manduca sexta Larvae. The Journal of biological chemistry, 282, 21222-21229.
http://dx.doi.org/10.1074/jbc.M701314200 [3] Alouf, J.E. and Popoff, M.R. (2006) The Comprehensive Sourcebook of Bacterial Protein Toxins. 3rd Edition, Elsevier, Amsterdam, Boston. [4] Schnepf, E., Crickmore, N., Van Rie, J., Lereclus, D., Baum, J., Feitelson, J., Zeigler, D.R. and Dean, D.H. (1998) Bacillus thuringiensis and Its Pesticidal Crystal Proteins. Microbiology and Molecular Biology Reviews: MMBR, 62, 775-806. [5] Knowles, B.H. (1994) Mechanism of Action of Bacillus thuringiensis Insecticidal δ-Endotoxins. Advances in Insect Physiology, 24, 275-308.
http://dx.doi.org/10.1016/s0065-2806(08)60085-5 [6] Dean, D.H., Rajamohan, F., Lee, M.K., Wu, S.J., Chen, X.J., Alcantara, E. and Hussain, S.R. (1996) Probing the Mechanism of Action of Bacillus thuringiensis Insecticidal Proteins by Site-Directed Mutagenesis—A Minireview. Gene, 179, 111-117.
http://dx.doi.org/10.1016/S0378-1119(96)00442-8 [7] Chattopadhyay, A., Bhatnagar, N.B. and Bhatnagar, R. (2004) Bacterial Insecticidal Toxins. Critical Reviews in Microbiology, 30, 33-54. [8] Soberon, M., Pardo, L., Munoz-Garay, C., Sanchez, J., Gomez, I., Porta, H. and Bravo, A. (2010) Pore Formation by Cry Toxins. Advances in Experimental Medicine and Biology, 677, 127-142.
http://dx.doi.org/10.1007/978-1-4419-6327-7_11 [9] Mueller, M., Grauschopf, U., Maier, T., Glockshuber, R. and Ban, N. (2009) The Structure of a Cytolytic Alpha-Helical Toxin Pore Reveals Its Assembly Mechanism. Nature, 459, 726-730.
http://dx.doi.org/10.1038/nature08026 [10] Shepard, L.A., Heuck, A.P., Hamman, B.D., Rossjohn, J., Parker, M.W., Ryan, K.R., Johnson, A.E. and Tweten, R.K. (1998) Identification of a Membrane-Spanning Domain of the Thiol-Activated Pore-Forming Toxin Clostridium perfringens Perfringolysin O: An Alpha-Helical to Beta-Sheet Transition Identified by Fluorescence Spectroscopy. Biochemistry, 37, 14563-14574.
http://dx.doi.org/10.1021/bi981452f [11] Collier, R.J. and Young, J.A.T. (2003) Anthrax Toxin. Annual Review of Cell and Developmental Biology, 19, 45-70.
http://dx.doi.org/10.1146/annurev.cellbio.19.111301.140655 [12] Young, J.A. and Collier, R.J. (2007) Anthrax Toxin: Receptor Binding, Internalization, Pore Formation, and Translocation. Annual Review of Biochemistry, 76, 243-265.
http://dx.doi.org/10.1146/annurev.biochem.75.103004.142728 [13] Tilley, S.J., Orlova, E.V., Gilbert, R.J., Andrew, P.W. and Saibil, H.R. (2005) Structural Basis of Pore Formation by the Bacterial Toxin Pneumolysin. Cell, 121, 247-256.
http://dx.doi.org/10.1016/j.cell.2005.02.033 [14] Aronson, A.I., Geng, C. and Wu, L. (1999) Aggregation of Bacillus thuringiensis Cry1A Toxins upon Binding to Target Insect Larval Midgut Vesicles. Applied and Environmental Microbiology, 65, 2503-2507. [15] Tomimoto, K., Hayakawa, T. and Hori, H. (2006) Pronase Digestion of Brush Border Membrane-Bound Cry1Aa Shows That Almost the Whole Activated Cry1Aa Molecule Penetrates into the Membrane. Comparative Biochemistry and Physiology Part B: Biochemistry & Molecular Biology, 144, 413-422.
http://dx.doi.org/10.1016/j.cbpb.2006.04.013 [16] Bravo, A., Gomez, I., Conde, J., Munoz-Garay, C., Sanchez, J., Miranda, R., Zhuang, M., Gill, S.S. and Soberon, 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, 1667, 38-46.
http://dx.doi.org/10.1016/j.bbamem.2004.08.013 [17] Rausell, C., Munoz-Garay, C., Miranda-CassoLuengo, R., Gomez, I., Rudino-Pinera, E., Soberón, M. and Bravo, A. (2004) 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, 166-174.
http://dx.doi.org/10.1021/bi035527d [18] Gomez, I., Dean, D.H., Bravo, A. and Soberon, M. (2003) Molecular Basis for Bacillus thuringiensis Cry1Ab Toxin Specificity: Two Structural Determinants in the Manduca sexta Bt-R1 Receptor Interact with Loops Alpha-8 and 2 in Domain II of Cy1Ab Toxin. Biochemistry, 42, 10482-10489.
http://dx.doi.org/10.1021/bi034440p [19] Gomez, I., Miranda-Rios, J., Rudino-Pinera, E., Oltean, D.I., Gill, S.S., Bravo, A. and Soberon, M. (2002) Hydropathic Complementarity Determines Interaction of Epitope 869HITDTNNK876 in Manduca sexta Bt-R1 Receptor with Loop 2 of Domain II of Bacillus thuringiensis Cry1A Toxins. The Journal of Biological Chemistry, 277, 30137-30143.
http://dx.doi.org/10.1074/jbc.M203121200 [20] Pacheco, S., Gomez, I., Arenas, I., Saab-Rincon, G., Rodriguez-Almazan, C., Gill, S.S., Bravo, A. and Soberon, M. (2009) Domain II Loop 3 of Bacillus thuringiensis Cry1Ab Toxin Is Involved in a “Ping Pong” Binding Mechanism with Manduca sexta Aminopeptidase-N and Cadherin Receptors. The Journal of Biological Chemistry, 284, 32750-32757.
http://dx.doi.org/10.1074/jbc.M109.024968 [21] Arenas, I., Bravo, A., Soberon, M. and Gomez, I. (2010) Role of Al-kaline Phosphatase from Manduca sexta in the Mechanism of Action of Bacillus thuringiensis Cry1Ab Toxin. The Journal of Biological Chemistry, 285, 12497- 12503.
http://dx.doi.org/10.1074/jbc.M109.085266 [22] Lee, M.K., Milne, R.E., Ge, A.Z. and Dean, D.H. (1992) Location of a Bombyx mori Receptor Binding Region on a Bacillus thuringiensis Delta-Endotoxin. The Journal of Biological Chemistry, 267, 3115-3121. [23] Jimenez-Juarez, N., Munoz-Garay, C., Gomez, I., Gill, S.S., Soberon, M. and Bravo, A. (2008) The Pre-Pore from Bacillus thuringiensis Cry1Ab Toxin Is Necessary to Induce Insect Death in Manduca sexta. Peptides, 29, 318-323.
http://dx.doi.org/10.1016/j.peptides.2007.09.026 [24] Nair, M.S. and Dean, D.H. (2008) All Domains of Cry1A Toxins Insert into Insect Brush Border Membranes. The Journal of Biological Chemistry, 283, 26324-26331.
http://dx.doi.org/10.1074/jbc.M802895200 [25] Chen, J., Hua, G., Jurat-Fuentes, J.L., Abdullah, M.A. and Adang, M.J. (2007) Synergism of Bacillus thuringiensis Toxins by a Fragment of a Toxin-Binding Cadherin. Proceedings of the National Academy of Sciences of the United States of America, 104, 13901-13906.
http://dx.doi.org/10.1073/pnas.0706011104 [26] Zhang, X., Candas, M., Griko, N.B., Taussig, R. and Bulla Jr., L.A. (2006) A Mechanism of Cell Death Involving an Adenylyl Cyclase/PKA Signaling Pathway Is Induced by the Cry1Ab Toxin of Bacillus thuringiensis. Proceedings of the National Academy of Sciences of the United States of America, 103, 9897-9902.
http://dx.doi.org/10.1073/pnas.0604017103 [27] Griko, N., Zhang, X., Ibrahim, M., Midboe, E.G. and Bulla Jr., L.A. (2008) Susceptibility of Manduca sexta to Cry1Ab Toxin of Bacillus thuringiensis Correlates Directly to Developmental Expression of the Cadherin Receptor BT-R1. Comparative Biochemistry and Physiology Part B: Biochemistry & Molecular Biology, 151, 59-63.
http://dx.doi.org/10.1016/j.cbpb.2008.05.016
[1] Gomez, I., Sanchez, J., Miranda, R., Bravo, A. and Soberon, M. (2002) Cadherin-Like Receptor Binding Facilitates Proteolytic Cleavage of Helix Alpha-1 in Domain I and Oligomer Pre-Pore Formation of Bacillus thuringiensis Cry1Ab Toxin. FEBS Letters, 513, 242-246.
http://dx.doi.org/10.1016/S0014-5793(02)02321-9 [2] Jimenez-Juarez, N., Munoz-Garay, C., Gomez, I., Saab-Rincon, G., Damian-Almazo, J.Y., Gill, S.S., Soberon, M. and Bravo, A. (2007) Bacillus thuringiensis Cry1Ab Mutants Affecting Oligomer Formation Are Non-Toxic to Manduca sexta Larvae. The Journal of biological chemistry, 282, 21222-21229.
http://dx.doi.org/10.1074/jbc.M701314200 [3] Alouf, J.E. and Popoff, M.R. (2006) The Comprehensive Sourcebook of Bacterial Protein Toxins. 3rd Edition, Elsevier, Amsterdam, Boston. [4] Schnepf, E., Crickmore, N., Van Rie, J., Lereclus, D., Baum, J., Feitelson, J., Zeigler, D.R. and Dean, D.H. (1998) Bacillus thuringiensis and Its Pesticidal Crystal Proteins. Microbiology and Molecular Biology Reviews: MMBR, 62, 775-806. [5] Knowles, B.H. (1994) Mechanism of Action of Bacillus thuringiensis Insecticidal δ-Endotoxins. Advances in Insect Physiology, 24, 275-308.
http://dx.doi.org/10.1016/s0065-2806(08)60085-5 [6] Dean, D.H., Rajamohan, F., Lee, M.K., Wu, S.J., Chen, X.J., Alcantara, E. and Hussain, S.R. (1996) Probing the Mechanism of Action of Bacillus thuringiensis Insecticidal Proteins by Site-Directed Mutagenesis—A Minireview. Gene, 179, 111-117.
http://dx.doi.org/10.1016/S0378-1119(96)00442-8 [7] Chattopadhyay, A., Bhatnagar, N.B. and Bhatnagar, R. (2004) Bacterial Insecticidal Toxins. Critical Reviews in Microbiology, 30, 33-54. [8] Soberon, M., Pardo, L., Munoz-Garay, C., Sanchez, J., Gomez, I., Porta, H. and Bravo, A. (2010) Pore Formation by Cry Toxins. Advances in Experimental Medicine and Biology, 677, 127-142.
http://dx.doi.org/10.1007/978-1-4419-6327-7_11 [9] Mueller, M., Grauschopf, U., Maier, T., Glockshuber, R. and Ban, N. (2009) The Structure of a Cytolytic Alpha-Helical Toxin Pore Reveals Its Assembly Mechanism. Nature, 459, 726-730.
http://dx.doi.org/10.1038/nature08026 [10] Shepard, L.A., Heuck, A.P., Hamman, B.D., Rossjohn, J., Parker, M.W., Ryan, K.R., Johnson, A.E. and Tweten, R.K. (1998) Identification of a Membrane-Spanning Domain of the Thiol-Activated Pore-Forming Toxin Clostridium perfringens Perfringolysin O: An Alpha-Helical to Beta-Sheet Transition Identified by Fluorescence Spectroscopy. Biochemistry, 37, 14563-14574.
http://dx.doi.org/10.1021/bi981452f [11] Collier, R.J. and Young, J.A.T. (2003) Anthrax Toxin. Annual Review of Cell and Developmental Biology, 19, 45-70.
http://dx.doi.org/10.1146/annurev.cellbio.19.111301.140655 [12] Young, J.A. and Collier, R.J. (2007) Anthrax Toxin: Receptor Binding, Internalization, Pore Formation, and Translocation. Annual Review of Biochemistry, 76, 243-265.
http://dx.doi.org/10.1146/annurev.biochem.75.103004.142728 [13] Tilley, S.J., Orlova, E.V., Gilbert, R.J., Andrew, P.W. and Saibil, H.R. (2005) Structural Basis of Pore Formation by the Bacterial Toxin Pneumolysin. Cell, 121, 247-256.
http://dx.doi.org/10.1016/j.cell.2005.02.033 [14] Aronson, A.I., Geng, C. and Wu, L. (1999) Aggregation of Bacillus thuringiensis Cry1A Toxins upon Binding to Target Insect Larval Midgut Vesicles. Applied and Environmental Microbiology, 65, 2503-2507. [15] Tomimoto, K., Hayakawa, T. and Hori, H. (2006) Pronase Digestion of Brush Border Membrane-Bound Cry1Aa Shows That Almost the Whole Activated Cry1Aa Molecule Penetrates into the Membrane. Comparative Biochemistry and Physiology Part B: Biochemistry & Molecular Biology, 144, 413-422.
http://dx.doi.org/10.1016/j.cbpb.2006.04.013 [16] Bravo, A., Gomez, I., Conde, J., Munoz-Garay, C., Sanchez, J., Miranda, R., Zhuang, M., Gill, S.S. and Soberon, 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, 1667, 38-46.
http://dx.doi.org/10.1016/j.bbamem.2004.08.013 [17] Rausell, C., Munoz-Garay, C., Miranda-CassoLuengo, R., Gomez, I., Rudino-Pinera, E., Soberón, M. and Bravo, A. (2004) 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, 166-174.
http://dx.doi.org/10.1021/bi035527d [18] Gomez, I., Dean, D.H., Bravo, A. and Soberon, M. (2003) Molecular Basis for Bacillus thuringiensis Cry1Ab Toxin Specificity: Two Structural Determinants in the Manduca sexta Bt-R1 Receptor Interact with Loops Alpha-8 and 2 in Domain II of Cy1Ab Toxin. Biochemistry, 42, 10482-10489.
http://dx.doi.org/10.1021/bi034440p [19] Gomez, I., Miranda-Rios, J., Rudino-Pinera, E., Oltean, D.I., Gill, S.S., Bravo, A. and Soberon, M. (2002) Hydropathic Complementarity Determines Interaction of Epitope 869HITDTNNK876 in Manduca sexta Bt-R1 Receptor with Loop 2 of Domain II of Bacillus thuringiensis Cry1A Toxins. The Journal of Biological Chemistry, 277, 30137-30143.
http://dx.doi.org/10.1074/jbc.M203121200 [20] Pacheco, S., Gomez, I., Arenas, I., Saab-Rincon, G., Rodriguez-Almazan, C., Gill, S.S., Bravo, A. and Soberon, M. (2009) Domain II Loop 3 of Bacillus thuringiensis Cry1Ab Toxin Is Involved in a “Ping Pong” Binding Mechanism with Manduca sexta Aminopeptidase-N and Cadherin Receptors. The Journal of Biological Chemistry, 284, 32750-32757.
http://dx.doi.org/10.1074/jbc.M109.024968 [21] Arenas, I., Bravo, A., Soberon, M. and Gomez, I. (2010) Role of Al-kaline Phosphatase from Manduca sexta in the Mechanism of Action of Bacillus thuringiensis Cry1Ab Toxin. The Journal of Biological Chemistry, 285, 12497- 12503.
http://dx.doi.org/10.1074/jbc.M109.085266 [22] Lee, M.K., Milne, R.E., Ge, A.Z. and Dean, D.H. (1992) Location of a Bombyx mori Receptor Binding Region on a Bacillus thuringiensis Delta-Endotoxin. The Journal of Biological Chemistry, 267, 3115-3121. [23] Jimenez-Juarez, N., Munoz-Garay, C., Gomez, I., Gill, S.S., Soberon, M. and Bravo, A. (2008) The Pre-Pore from Bacillus thuringiensis Cry1Ab Toxin Is Necessary to Induce Insect Death in Manduca sexta. Peptides, 29, 318-323.
http://dx.doi.org/10.1016/j.peptides.2007.09.026 [24] Nair, M.S. and Dean, D.H. (2008) All Domains of Cry1A Toxins Insert into Insect Brush Border Membranes. The Journal of Biological Chemistry, 283, 26324-26331.
http://dx.doi.org/10.1074/jbc.M802895200 [25] Chen, J., Hua, G., Jurat-Fuentes, J.L., Abdullah, M.A. and Adang, M.J. (2007) Synergism of Bacillus thuringiensis Toxins by a Fragment of a Toxin-Binding Cadherin. Proceedings of the National Academy of Sciences of the United States of America, 104, 13901-13906.
http://dx.doi.org/10.1073/pnas.0706011104 [26] Zhang, X., Candas, M., Griko, N.B., Taussig, R. and Bulla Jr., L.A. (2006) A Mechanism of Cell Death Involving an Adenylyl Cyclase/PKA Signaling Pathway Is Induced by the Cry1Ab Toxin of Bacillus thuringiensis. Proceedings of the National Academy of Sciences of the United States of America, 103, 9897-9902.
http://dx.doi.org/10.1073/pnas.0604017103 [27] Griko, N., Zhang, X., Ibrahim, M., Midboe, E.G. and Bulla Jr., L.A. (2008) Susceptibility of Manduca sexta to Cry1Ab Toxin of Bacillus thuringiensis Correlates Directly to Developmental Expression of the Cadherin Receptor BT-R1. Comparative Biochemistry and Physiology Part B: Biochemistry & Molecular Biology, 151, 59-63.
http://dx.doi.org/10.1016/j.cbpb.2008.05.016