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Repurposing Bacterial Toxins
Bacterial toxins can be repurposed. These diverse macromolecular complexes bind to human cells and transport enzymes across membranes. Some are currently used in medical clinics without extensive biomolecular engineering. Biomedical and biotechnological research is yielding information and techniques that allows for the design of repurposed toxins by “shuffling” the components of AB-type binary exotoxins toxins. These engineered proteins can be applied to a breadth of biomedical applications. Here, the Clostridium botulinum C2 toxin enzymatic and translocation components were combined with the neurological targeting function of the C. botulinum serotype C1 binding component. 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. In subsequent work, the C. botulinum C2 toxin binding domain was independently produced in a preliminary effort to engineer binding functions of a series of repurposed toxins, or “R2Cs”. Future work will evaluate the electrophysiological effects of mutagenized R2Cs in the context of nerve signaling. This may result in an enhanced approach to diagnostics of peripheral neuropathies by identification of characteristic neuropathic signatures.
Pavlik, Benjamin J, "Repurposing Bacterial Toxins" (2017). ETD collection for University of Nebraska - Lincoln. AAI10682737.