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Retargeting the Clostridium botulinum C2 toxin to the neuronal cytosol
Many biological toxins are known to attack specific cell types, delivering their enzymatic payloads to the cytosol. This process can be manipulated by molecular engineering of chimeric toxins. Using toxins with naturally unlinked components as a starting point is advantageous because it allows for t...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Nature Publishing Group
2016
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4812341/ https://www.ncbi.nlm.nih.gov/pubmed/27025362 http://dx.doi.org/10.1038/srep23707 |
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author | Pavlik, Benjamin J. Hruska, Elizabeth J. Van Cott, Kevin E. Blum, Paul H. |
author_facet | Pavlik, Benjamin J. Hruska, Elizabeth J. Van Cott, Kevin E. Blum, Paul H. |
author_sort | Pavlik, Benjamin J. |
collection | PubMed |
description | Many biological toxins are known to attack specific cell types, delivering their enzymatic payloads to the cytosol. This process can be manipulated by molecular engineering of chimeric toxins. Using toxins with naturally unlinked components as a starting point is advantageous because it allows for the development of payloads separately from the binding/translocation components. Here the Clostridium botulinum C2 binding/translocation domain was retargeted to neural cell populations by deleting its non-specific binding domain and replacing it with a C. botulinum neurotoxin binding domain. This fusion protein was used to deliver fluorescently labeled payloads to Neuro-2a cells. Intracellular delivery was quantified by flow cytometry and found to be dependent on artificial enrichment of cells with the polysialoganglioside receptor GT1b. Visualization by confocal microscopy showed a dissociation of payloads from the early endosome indicating translocation of the chimeric toxin. The natural Clostridium botulinum C2 toxin was then delivered to human glioblastoma A172 and synchronized HeLa cells. In the presence of the fusion protein, native cytosolic enzymatic activity of the enzyme was observed and found to be GT1b-dependent. This retargeted toxin may enable delivery of therapeutics to peripheral neurons and be of use in addressing experimental questions about neural physiology. |
format | Online Article Text |
id | pubmed-4812341 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2016 |
publisher | Nature Publishing Group |
record_format | MEDLINE/PubMed |
spelling | pubmed-48123412016-04-04 Retargeting the Clostridium botulinum C2 toxin to the neuronal cytosol Pavlik, Benjamin J. Hruska, Elizabeth J. Van Cott, Kevin E. Blum, Paul H. Sci Rep Article Many biological toxins are known to attack specific cell types, delivering their enzymatic payloads to the cytosol. This process can be manipulated by molecular engineering of chimeric toxins. Using toxins with naturally unlinked components as a starting point is advantageous because it allows for the development of payloads separately from the binding/translocation components. Here the Clostridium botulinum C2 binding/translocation domain was retargeted to neural cell populations by deleting its non-specific binding domain and replacing it with a C. botulinum neurotoxin binding domain. This fusion protein was used to deliver fluorescently labeled payloads to Neuro-2a cells. Intracellular delivery was quantified by flow cytometry and found to be dependent on artificial enrichment of cells with the polysialoganglioside receptor GT1b. Visualization by confocal microscopy showed a dissociation of payloads from the early endosome indicating translocation of the chimeric toxin. The natural Clostridium botulinum C2 toxin was then delivered to human glioblastoma A172 and synchronized HeLa cells. In the presence of the fusion protein, native cytosolic enzymatic activity of the enzyme was observed and found to be GT1b-dependent. This retargeted toxin may enable delivery of therapeutics to peripheral neurons and be of use in addressing experimental questions about neural physiology. Nature Publishing Group 2016-03-30 /pmc/articles/PMC4812341/ /pubmed/27025362 http://dx.doi.org/10.1038/srep23707 Text en Copyright © 2016, Macmillan Publishers Limited http://creativecommons.org/licenses/by/4.0/ This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/ |
spellingShingle | Article Pavlik, Benjamin J. Hruska, Elizabeth J. Van Cott, Kevin E. Blum, Paul H. Retargeting the Clostridium botulinum C2 toxin to the neuronal cytosol |
title | Retargeting the Clostridium botulinum C2 toxin to the neuronal cytosol |
title_full | Retargeting the Clostridium botulinum C2 toxin to the neuronal cytosol |
title_fullStr | Retargeting the Clostridium botulinum C2 toxin to the neuronal cytosol |
title_full_unstemmed | Retargeting the Clostridium botulinum C2 toxin to the neuronal cytosol |
title_short | Retargeting the Clostridium botulinum C2 toxin to the neuronal cytosol |
title_sort | retargeting the clostridium botulinum c2 toxin to the neuronal cytosol |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4812341/ https://www.ncbi.nlm.nih.gov/pubmed/27025362 http://dx.doi.org/10.1038/srep23707 |
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