Single Molecule Detection of Intermediates During Botulinum Neurotoxin Translocation Across Membranes.
From: Section of Neurobiology, Division of Biological Sciences, University of California at San Diego, La Jolla, CA 92093-0366, USA.
Proceedings of the National Academy of Sciences of the United States of America
- Publish Date: Jun 2007
- ISSN: 0027-8424
- Volume: 104
- Issue: 25
- Pages: 10447-52
- Medium: Print
- Language: English
- Citation (JAMA): Fischer Audrey, Montal Mauricio, et al. Single Molecule Detection of Intermediates During Botulinum Neurotoxin Translocation Across Membranes.. Proc. Natl. Acad. Sci. U.S.A. Jun 2007;104:10447-52
Abstract
The dynamics of Clostridium botulinum neurotoxins (BoNTs) protein-translocation across membranes was investigated by using a single molecule assay with millisecond resolution on excised patches of neuronal cells. Translocation of BoNT/A light chain (LC) by heavy chain (HC) was observed in real time as an increase of channel conductance: the HC channel is occluded by the LC during transit, then unoccluded after completion of translocation and release of LC-cargo. We identified an entirely unknown succession of intermediate conductance stages during LC translocation. For the single-chain BoNT/E, by contrast to the di-chain BoNT/A, we demonstrate that productive translocation requires proteolysis of the LC cargo from the HC chaperone. We propose a model for the set of protein-protein interactions between translocase and cargo at each step of translocation that supports the notion of an interdependent, tight interplay between the HC chaperone and the LC cargo preventing LC aggregation and dictating the outcome of translocation: productive passage of cargo or abortive channel occlusion by cargo.
Mesh Headings (Keywords): Animals, Biological Assay, Botulinum Toxin Type A, Botulinum Toxins, Cell Line, Tumor, Cell Membrane, Clostridium botulinum, Disulfides, Hydrogen-Ion Concentration, Hydrolysis, Immunoglobulin Fab Fragments, Mice, Models, Biological, Molecular Chaperones, Neuroblastoma, Neurons, Neurotoxins, Oxidation-Reduction, Patch-Clamp Techniques, Protein Denaturation, Protein Isoforms, Protein Renaturation, Protein Transport, Sensitivity and Specificity, Time Factors
Check for Full Text / PubMed Unique Identifier (PMID): 17563359
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