| Genetically engineered clostridial C2 toxin as a novel delivery system for living mammalian cells. | |
| | |
MedLine Citation:
|
PMID: 20030334 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
|
The C2 toxin of Clostridium botulinum is a binary bacterial protein toxin, comprising the enzyme component C2I and the separate binding/translocation component C2IIa. C2IIa mediates the transport of C2I into the host cell cytosol. The N-terminal domain of C2I (C2IN) is enzymatically inactive but essential for C2IIa-mediated internalization of C2I. Here, we exploit the C2IIa/C2IN system to generate a recombinant C2IN-streptavidin fusion protein allowing for the delivery of biotinylated molecules into the cytosol of mammalian cells. C2IN-streptavidin overproduced in E. coli was affinity-purified and capable of binding biotinylated proteins in a concentration-dependent manner. Real-time surface plasmon resonance confirmed the biotin-mediated interaction yielding a K(D)-value of approximately 0.75 muM. Internalization of C2IN-streptavidin into the cytosol of epithelial cells and macrophages was demonstrated by immunoblot analysis and confirmed by confocal microscopy. Cell viability studies showed no cytotoxic effects of the novel transporter. Furthermore, Vero cells treated with biotin-fluorescein or biocytin-Alexa488 as model cargo displayed a specific C2IN-streptavidin/C2IIa-dependent uptake, providing proof-of-principle for the functionality of this novel delivery system. |
| | |
Authors:
|
J?rg Fahrer; Rainer Plunien; Ulrike Binder; Torben Langer; Hartmut Seliger; Holger Barth |
Related Documents
:
|
2171494 - Cyclic amp accumulation in hela cells induced by cholera toxin. involvement of the cera... 11445574 - Comparative biochemical and immunocytochemical studies reveal differences in the effect... 7956944 - Gonadotropin-releasing hormone (gnrh)-receptor coupling to inositol phosphate and prola... 11182834 - Pathomorphological characteristics of experimental toxemia induced by thermostable yers... 2918314 - Effect of glucose starvation on glucose transport in neuronal cells in primary culture ... 9496734 - Dipyridamole inhibits human mesangial cell proliferation. |
Publication Detail:
|
Type: Journal Article |
Journal Detail:
|
Title: Bioconjugate chemistry Volume: 21 ISSN: 1520-4812 ISO Abbreviation: Bioconjug. Chem. Publication Date: 2010 Jan |
Date Detail:
|
Created Date: 2010-01-20 Completed Date: 2010-05-20 Revised Date: - |
Medline Journal Info:
|
Nlm Unique ID: 9010319 Medline TA: Bioconjug Chem Country: United States |
Other Details:
|
Languages: eng Pagination: 130-9 Citation Subset: IM |
Affiliation:
|
Institute of Pharmacology and Toxicology, University of Ulm, Albert-Einstein-Allee 47, D-89081 Ulm, Germany. |
Export Citation:
|
APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
|
Animals Binding Sites Biotin / chemistry, metabolism Biotinylation Botulinum Toxins / chemistry, genetics*, metabolism*, toxicity Cell Line Cell Survival Cercopithecus aethiops Cricetinae Cytosol / metabolism Drug Delivery Systems / methods* Epithelial Cells / metabolism Escherichia coli / chemistry, metabolism Fluoresceins / chemistry, metabolism Genetic Engineering* Humans Immunoblotting Macrophages / metabolism Microscopy, Confocal Protein Transport Recombinant Fusion Proteins / chemistry, genetics*, metabolism*, toxicity Streptavidin / chemistry, genetics, metabolism Vero Cells |
| Chemical | |
Reg. No./Substance:
|
0/Botulinum Toxins; 0/Fluoresceins; 0/Recombinant Fusion Proteins; 58-85-5/Biotin; 9013-20-1/Streptavidin |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
Previous Document: Finding Out Egyptian Gods' Secret Using Analytical Chemistry: Biomedical Properties of Egyptian Blac...
Next Document: Photoelectrochemical Characterization of a Robust TiO(2)/BDD Heterojunction Electrode for Sensing Ap...