Interaction of Clostridium perfringens delta toxin with erythrocyte and liposome membranes and relation with the specific binding to the ganglioside GM2.
The specific interaction of the cytolytic Clostridium perfringens delta toxin with membrane GM2 was indicated by: (i) characterization of this glycolipid in the membrane of sheep and goat erythrocytes, which are lysed by the toxin, whereas GM2 was undetectable in insensitive rabbit erythrocytes, (ii) demonstration of 125I-toxin binding to GM2, by autoradiography, following incubation with thin-layer chromatograms containing separated neuroblastoma gangliosides, and (iii) toxin fixation by phospholipid-cholesterol unilamellar vesicles containing either sheep gangliosides or GM2. In order to investigate the intramembrane events leading to membrane disruption following toxin binding, the photoreactive probe 12(4-azido-2-nitrophenoxy)stearoyl 1-14C glucosamine, which inserts into the outer layer and labels integral membrane proteins, was used to establish whether delta toxin penetrates into target cell membrane. No toxin labeling was found, suggesting that toxin action takes place at the membrane surface. This contention is supported by the observation that despite toxin binding, GM2 liposomes did not release entrapped 14C-glucose. Treatment of toxin with carboxypeptidases, but not aminopeptidases, abolished both toxin binding capacity onto erythrocytes and its combination with antitoxin neutralizing antibodies, suggesting that the carboxy terminal end of the toxin is critical for binding to cell membrane.
Jolivet-Reynaud
C
C
Unité des Antigènes Bactériens, (UA CNRS 557), Institut Pasteur, Paris, France.
Hauttecoeur
B
B
Alouf
J E
JE
D000626:Aminopeptidases / Q000494:pharmacology; D000818:Animals; D001427:Bacterial Toxins / Q000378:metabolism / Q000494:pharmacology*; D001667:Binding, Competitive; D002268:Carboxypeptidases / Q000494:pharmacology; D002462:Cell Membrane / Q000187:drug effects*; D002855:Chromatography, Thin Layer; D003016:Clostridium perfringens / Q000472:pathogenicity; D004910:Erythrocyte Membrane / Q000187:drug effects; D005678:G(M2) Ganglioside / Q000378:metabolism*; D005732:Gangliosides / Q000097:blood / Q000378:metabolism*; D006041:Goats; D066298:In Vitro Techniques; D008081:Liposomes; D011817:Rabbits; D012756:Sheep
D001427:Bacterial Toxins; C029352:Clostridium perfringens delta-toxin; D005732:Gangliosides; D008081:Liposomes; D005678:G(M2) Ganglioside; D002268:Carboxypeptidases; D000626:Aminopeptidases