Studies on the Role of a Nucleoside‐Phosphate‐Binding Site of Diphtheria Toxin in the Binding of Toxin to Vero Cells or Liposomes

Publisher Website
Google Scholar
Abstract
Using Fab antibody fragments directed against protein crm 45 (non-toxic, cross-reacting material serologically-related to diphtheria toxin mutant protein, the product of a nonsense mutation in the diphtheria toxin gene) or the B 45 domain of this molecule (the remaining portion of the toxin B fragment present in protein crm 45), the diphtheria toxin polyphosphate-binding site (P site) evidenced by the ability of the toxin to bind ATP, was shown to be affected upon incubation of this protein with the Fab fragment of anti-protein (crm 45) IgG but not with the Fab fragment of anti-polypeptide (B 45) IgG. After incubation with an excess of the Fab fragment of anti-polypeptide (B 45) IgG, 125I-labeled toxin does not bind to toxin-sensitive Vero cells. Thus, the P site is probably not by itself the binding domain of diphtheria toxin to the cell membrane receptor; the region of the toxin molecule devoted to this particular activity overlapped probably on the B-45 polypeptide. With or without a functional P site, diphtheria toxin is able to bind liposomes. However, interaction with phospholipid vesicles with toxin is blocked upon incubation of the protein with the Fab fragment of anti-polypeptide (B 45) IgG. Thus, the P site of toxin evidently does not participate in the ability of this molecule to bind liposomes.