Evidence that membrane phospholipids and protein are required for binding of diphtheria toxin in Vero cells

Treatment with phospholipase C strongly protected monkey kidney (Vero) cells against diphtheria toxin and reduced the ability of the cells to bind 125I-labelled toxin. Treatment with phospholipase D and with trypsin also protected the cells, although to a lesser extent. Phospholipase A2 had no protective effect. Phospholipase C also protected fetal hamster kidney cells against the toxin. After removal of the enzymes, as well as after treatment of the cells with 4-acetamide 4'-isothiocyanostilbene 2,2'-disulfonic acid, diphtheria toxin binding capability was restored slowly, apparently by a process requiring protein synthesis, since cycloheximide blocked the restoration. The data indicate that both phospholipids and protein are involved in the binding sites for diphtheria toxin.