title: Evidence for direct insertion of fragments A and B of diphtheria toxin into model membranes.
abstract: The entry of diphtheria toxin into model membranes was studied using a membrane-restricted photoprobe to monitor insertion. The results provided direct evidence that the A and B domains of diphtheria toxin can both insert into such membranes. Optimal binding was achieved with negatively charged liposomes at pH 3.6. Under these conditions, the A and B domains of nicked as well as unnicked toxin inserted. At 0 degrees C, only fragment B inserted although toxin was still optimally bound. At neutral pH, there was little binding of toxin, and fragment B inserted preferentially. Brief exposure of the toxin to pH 3.6 followed by adjustment of the pH to 7 and subsequent incubation of the toxin with vesicles at neutral pH greatly facilitated toxin binding as well as insertion of both A and B domains, indicating also that a pH gradient was not required for these processes. Data obtained from the tryptic fragmentation patterns and circular dichroism spectra indicated that exposure to acidic pH had induced a conformational change in the toxin which may have exposed hydrophobic regions. A similar conformational change may occur in vivo after acidification of cytoplasmic vesicles into which toxin is delivered by receptor-mediated endocytosis. This could facilitate insertion of toxin into the vesicle membranes and subsequent translocation of fragment A to the cytosol, where it causes inhibition of protein synthesis.
mesh_terms: D002942:Circular Dichroism; D004167:Diphtheria Toxin / Q000032:analysis*; D006863:Hydrogen-Ion Concentration; D008081:Liposomes / Q000032:analysis; D008567:Membranes, Artificial; D010446:Peptide Fragments / Q000032:analysis*; D011487:Protein Conformation; D013696:Temperature; D014357:Trypsin / Q000378:metabolism
chemical_list: D004167:Diphtheria Toxin; D008081:Liposomes; D008567:Membranes, Artificial; D010446:Peptide Fragments; C029795:diphtheria toxin fragment A; C039171:diphtheria toxin fragment B; D014357:Trypsin