title: A conserved tetrameric interaction of cry toxin helix α3 suggests a functional role for toxin oligomerization.
abstract: Crystal (Cry) toxins are widely used for insect control, but their mechanism of toxicity is still uncertain. These toxins can form lytic pores in vitro, and water soluble tetrameric pre-pore intermediates have been reported. Even the precise oligomeric state of the toxin in membranes, trimeric or tetrameric, is still a debated issue. Based on previous reports, we have assumed that interactions between toxin monomers in solution are at least partly mediated by domain I, and we have analyzed in silico the homo-oligomerization tendencies of the domain I α-helices individually. Using many homologous sequences for each α-helix, our strategy allows selection of evolutionarily conserved interactions. These interactions appeared only in helices α3 and α5, but only α3 produced a suitably oriented or α-helical sample in lipid bilayers, forming homotetramers in C14-betaine, and allowing determination of its rotational orientation in lipid bilayers using site-specific infrared dichroism (SSID). The determined orientation in the tetrameric model is in agreement with only one of the evolutionarily conserved models. In addition mutation R99E, which was found to inhibit oligomerization experimentally, greatly destabilized the tetramer in molecular dynamic simulations. In this model, helix 3 is able to form inter-monomer interactions without significant rearrangements of domain I, which is compatible with the available crystal structure of Cry toxins in solution. The model presented here at least partially explains the reported tetrameric oligomerization of Cry toxins in solution and the inhibition of this oligomerization by a synthetic α3 peptide.
authors.affiliation: School of Biological Sciences, Nanyang Technological University, 60, Nanyang Drive, 637551, Singapore.
authors.forename: Xin
authors.initials: X
authors.lastname: Lin
authors.affiliation: School of Biological Sciences, Nanyang Technological University, 60, Nanyang Drive, 637551, Singapore.
authors.forename: Krupakar
authors.initials: K
authors.lastname: Parthasarathy
authors.affiliation: School of Biological Sciences, Nanyang Technological University, 60, Nanyang Drive, 637551, Singapore.
authors.forename: Wahyu
authors.initials: W
authors.lastname: Surya
authors.affiliation: School of Biological Sciences, Nanyang Technological University, 60, Nanyang Drive, 637551, Singapore.
authors.forename: Tong
authors.initials: T
authors.lastname: Zhang
authors.affiliation: School of Biological Sciences, Nanyang Technological University, 60, Nanyang Drive, 637551, Singapore.
authors.forename: Yuguang
authors.initials: Y
authors.lastname: Mu
authors.affiliation: School of Biological Sciences, Nanyang Technological University, 60, Nanyang Drive, 637551, Singapore. Electronic address: jtorres@ntu.edu.sg.
authors.forename: Jaume
authors.initials: J
authors.lastname: Torres
mesh_terms: D000595:Amino Acid Sequence; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000737:chemistry*; D002462:Cell Membrane / Q000737:chemistry; D004731:Endotoxins / Q000737:chemistry*; D006460:Hemolysin Proteins / Q000737:chemistry*; D007306:Insecticides / Q000737:chemistry*; D008051:Lipid Bilayers / Q000737:chemistry*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment
chemical_list: D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D004731:Endotoxins; D006460:Hemolysin Proteins; D007306:Insecticides; D008051:Lipid Bilayers; C030956:insecticidal crystal protein, Bacillus Thuringiensis
keywords: Coiled-coil; Cry toxin; Evolutionary conservation; Membrane insertion; Oligomerization