Direct Detection of Membrane-Inserting Fragments Defines the Translocation Pores of a Family of Pathogenic Toxins |
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Authors: | Kathleen E. Orrell Åsa Tellgren-Roth Mercedes Di Bernardo Zhifen Zhang Flavia Cuviello Jasmin Lundqvist Gunnar von Heijne IngMarie Nilsson Roman A. Melnyk |
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Affiliation: | 1. Molecular Medicine Program, The Hospital for Sick Children Research Institute, Toronto M5G 0A4, Ontario, Canada;2. Department of Biochemistry, University of Toronto, Toronto M5S 1A8, Ontario, Canada;3. Department of Biochemistry & Biophysics, Stockholm University, SE-106 91 Stockholm, Sweden |
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Abstract: | Large clostridial toxins (LCTs) are a family of homologous proteins toxins that are directly responsible for the symptoms associated with a number of clostridial infections that cause disease in humans and in other animals. LCTs damage tissues by delivering a glucosyltransferase domain, which inactivates small GTPases, across the endosomal membrane and into the cytosol of target cells. Elucidating the mechanism of translocation for LCTs has been hampered by difficulties associated with identifying marginally hydrophobic segments that insert into the bounding membrane to form the translocation pore. Here, we directly measured the membrane-insertion partitioning propensity for segments spanning the putative pore-forming region using a translocon-mediated insertion assay and synthetic peptides. We identified membrane-inserting segments, as well as a conserved and functionally important negatively charged residue that requires protonation for efficient membrane insertion. We provide a model of the LCT pore, which provides insights into translocation for this enigmatic family of α-helical translocases. |
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Keywords: | bacterial toxins large clostridial toxins membrane insertion translocation LCTs large clostridial toxins TM transmembrane TcdA and TcdB CD circular dichroism DPC dodecylphosphocholine DOPC Edited by James Bowie |
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