Open Access Dissertation
Date of Award
Andew L. Feig
Virulence factors of pathogenic bacteria are to be blamed for life-threatening infections such as diphtheria, anthrax, botulism, and tentanus. In the case of enzymatic exotoxins, disease arises from cytotoxic proteins, and cytotoxicity is acheived only after cell entry. This intrinsic mechanism for cell entry is intriguing from research and medical views. Along with a review on existing cargo delivery systems utilizing protein toxins and the usefulness of such a system, here is described the first reported Clostridium difficile toxin A fusion protein, luciferase-TcdA, and evidence of the successful transport of an active enzyme, luciferase, into the cytosol of vero cells. A feature that makes our system so attractive, is the auto-proteolytic event that releases the cargo protein after internalization. In addition to protein delivery, the exciting success of a peptide-based inhibitor is described. Almost complete inhibition of toxin A was observed in cellulo by a peptide, identified to reversibly bind to the active site, HQSPWHHGGGC, that was functionalized with an epoxy group, HQSPGepoxyHHGGGC. Placement of the functional group is crucial for toxin inhibition. Through studying protein toxins we not only gain insight necessary for defending against deadly diseases, but we can also learn to harness their distinctive properties for the development of novel biotechnology tools, such as a protein translocation system.
Kern, Stephanie Marie, "Development of a cargo delivery system and inhibition studies focused on clostridium difficile toxin a" (2012). Wayne State University Dissertations. 449.