Journal
JOURNAL OF MOLECULAR BIOLOGY
Volume 366, Issue 5, Pages 1558-1568Publisher
ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD
DOI: 10.1016/j.jmb.2006.12.057
Keywords
insecticidal toxin; Xenorhabdus nematophila; Bacillus thuringiensis; tetramer; cage-like structure
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Funding
- Wellcome Trust Funding Source: Medline
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A recently identified class of proteins conferring insecticidal activity to several bacteria within the Enterobacteriaceae family have potential for control of commercially important insect pests. Here, we report the first purification, biophysical characterisation. and 3-D structural analysis of one of the toxin components, XptA1, from Xenorhabdus nematophila PMF1296 to a resolution of 23 A. Membrane binding studies indicate that the three-component toxin system has a different mode of action from that of proteins from Bacillus thuringiensis (Bt). Biophysical characterisation of XptA1 suggests a mechanism of action of XptA1 whereby it first binds to the cell membrane forming a structure with a central cavity and forms a complex with its partners XptB1 and XptC1 producing the full insecticidal toxin. The structure of XptA1 is shown by a combination of electron microscopy, ultracentrifugation and circular dichroism spectroscopy to be a 1.15 MDa tetramer with a cage-like structure. Each of the four symmetry-related subunits has three well-defined domains and a longitudinal twist with one end narrower than the other. One third of the residues of XptA1 are alpha-helical and it is suggested the subunits associate partly via an alpha-helical coiled-coil interaction. XptA1 itself shows the same secondary structure at neutral pH and in an alkaline environment up to pH10.5. This pH tolerance indicates that the folded XptA1 can pass through the midgut of Lepidopteran insects susceptible to the insecticidal toxin complex. This implies therefore that its folded structure is important for its biological activity. (c) 2007 Elsevier Ltd. All rights reserved.
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