Investigation of non-toxic diphtheria toxin mutant for bioconjugation studies with self-assembling vesicle polymers.

Dominic J. Rucco, Amanda M. Pritzlaff, Daniel A. Savin

George and Josephine Butler Polymer Research Laboratory
Center for Macromolecular Science and Engineering
Department of Chemistry

In this work we seek to exploit the targeting and cell entry mechanisms of a non-toxic DT mutant by polymer conjugation to self-assembled vesicles in an effort to improve local drug delivery in cancer therapy.  Diphtheria toxin (DT) provides an example whereby complex and specific actions are achieved through well-orchestrated mechanisms taking advantage of distinguished protein domains.  In short, recognition of cell bound heparin-binding EGF-like growth factor (HB-EGF), receptor mediated endocytosis, endosomal pore formation through lipid soluble loops and helix insertion, ADP-ribosylation of eukaryotic elongation factor 2 (EEF2), and subsequent cell death all result from the receptor, translocation, and catalytic domains. Currently, we are developing a protocol based on literature procedures to maximize efficiency of production, bioconjucation and ultimate study of DT-polymer systems, and here we present the overexpression of a DT mutant from E. Coli for eventual polymer bioconjugation.