Diphtheria toxin:receptor interaction: association, dissociation, and effect of pH.

Diphtheria toxin (DT) binds to a specific heparin-binding epidermal growth factor-like growth factor (HB-EGF) precursor that is expressed in DT-sensitive cells. DT binds to the cell-surface HB-EGF precursor with an apparent dissociation constant (KD) of approximately 1 x 10(-8) - 10(-9) M at 4 degreesC, a temperature at which toxin binds but is not internalized. The interaction of DT with the cell-surface receptor, however, may be influenced by other cell-surface components. We used a biosensor method to measure the binding of DT to immobilized recombinant human HB-EGF (hHB-EGF) at 25 degreesC with no other cellular components present. We observed that at pH 7.4, using this in vitro two component system, DT binds to hHB-EGF with an apparent KD of 2.7 x 10(-8) M. We also observed that the dissociation of DT from hHB-EGF at pH values that approach those of the endosome occurs at a faster rate as the pH is decreased. These results suggest that the low pH of the endosome is sufficient to allow DT to dissociate from the HB-EGF precursor, prior to the translocation of the enzymatically active fragment of DT into the cytosol.