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Please use this identifier to cite or link to this item: http://repository.li.mahidol.ac.th/dspace/handle/123456789/35395
Title: Potential prepore trimer formation by the Bacillus thuringiensis mosquito-specific toxin: Molecular insights into a critical prerequisite of membrane-bound monomers
Authors: Wilaiwan Sriwimol
Aratee Aroonkesorn
Somsri Sakdee
Chalermpol Kanchanawarin
Takayuki Uchihashi
Toshio Ando
Chanan Angsuthanasombat
Mahidol University
Kasetsart University
Kanazawa University
Biophysics Institute for Research and Development
Keywords: Biochemistry, Genetics and Molecular Biology
Issue Date: 21-Aug-2015
Citation: Journal of Biological Chemistry. Vol.290, No.34 (2015), 20793-20803
Abstract: © 2015 by The American Society for Biochemistry and Molecular Biology, Inc. The insecticidal feature of the three-domain Cry δ-endotoxins from Bacillus thuringiensis is generally attributed to their capability to form oligomeric pores, causing lysis of target larval midgut cells. However, the molecular description of their oligomerization process has not been clearly defined. Here a stable prepore of the 65-kDa trypsin-activated Cry4Ba mosquito-specific toxin was established through membrane-mimetic environments by forming an ∼200-kDa octyl-β-D-glucoside micelle-induced trimer. The SDS-resistant trimer caused cytolysis to Sf9 insect cells expressing Aedes-mALP (a Cry4Ba receptor) and was more effective than a toxin monomer in membrane perturbation of calcein-loaded liposomes. A three-dimensional model of toxin trimer obtained by negative-stain EM in combination with single-particle reconstruction at ∼5 nm resolution showed a propeller-shaped structure with 3-fold symmetry. Fitting the three-dimensional reconstructed EM map with a 100-ns molecular dynamics-simulated Cry4Ba structure interacting with an octyl-β-D-glucoside micelle showed relative positioning of individual domains in the context of the trimeric complex with a major protrusion from the pore-forming domain. Moreover, high-speed atomic force microscopy imaging at nanometer resolution and a subsecond frame rate demonstrated conformational transitions from a propeller-like to a globularly shaped trimer upon lipid membrane interactions, implying prepore-to-pore conversion. Real-time trimeric arrangement of monomers associated with L-α-dimyristoylphosphatidyl-choline/3-[(3-cholamidopropyl)dimethylammonio]-2-hydroxy-1-propanesulfonic acid bicelle membranes was also envisaged by successive high-speed atomic force microscopy imaging, depicting interactions among three individual subunits toward trimer formation. Together, our data provide the first pivotal insights into the structural requirement of membrane-induced conformational changes of Cry4Ba toxin monomers for the molecular assembly of a prepore trimer capable of inserting into target membranes to generate a lytic pore.
URI: https://www.scopus.com/inward/record.uri?partnerID=HzOxMe3b&scp=84939832278&origin=inward
http://repository.li.mahidol.ac.th/dspace/handle/123456789/35395
ISSN: 1083351X
00219258
Appears in Collections:Scopus 2011-2015

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