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dc.contributor.authorWilaiwan Sriwimolen_US
dc.contributor.authorAratee Aroonkesornen_US
dc.contributor.authorSomsri Sakdeeen_US
dc.contributor.authorChalermpol Kanchanawarinen_US
dc.contributor.authorTakayuki Uchihashien_US
dc.contributor.authorToshio Andoen_US
dc.contributor.authorChanan Angsuthanasombaten_US
dc.contributor.otherMahidol Universityen_US
dc.contributor.otherKasetsart Universityen_US
dc.contributor.otherKanazawa Universityen_US
dc.contributor.otherBiophysics Institute for Research and Developmenten_US
dc.identifier.citationJournal of Biological Chemistry. Vol.290, No.34 (2015), 20793-20803en_US
dc.description.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.en_US
dc.rightsMahidol Universityen_US
dc.subjectBiochemistry, Genetics and Molecular Biologyen_US
dc.titlePotential prepore trimer formation by the Bacillus thuringiensis mosquito-specific toxin: Molecular insights into a critical prerequisite of membrane-bound monomersen_US
Appears in Collections:Scopus 2011-2015

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