The C terminus of component C2II of Clostridium botulinum C2 toxin is essential for receptor binding

The binary Clostridium botulinum C2 toxin consists of two separate proteins , the binding component C2II (80.5 kDa) and the actin-ADP-ribosylating enzy me component C2I (49.4 kDa). For its cytotoxic action, C2II binds to a cell membrane receptor and induces cell entry of C2I via receptor-mediated endo cytosis. Here we studied the structure-function relationship of C2II by con structing truncated C2II proteins and producing polyclonal antisera against selective regions of C2II. An antibody raised against the C terminus (amin o acids 592 to 721) of C2II inhibited binding of C2II to cells. The antibod y prevented pore formation by C2II oligomers in artificial membranes but di d not influence the properties of existing channels. To further define the region responsible for receptor binding, we constructed proteins with delet ions in C2II; specifically, they lacked amino acid residues 592 to 721 and the 7 C-terminal amino acid residues. The truncated proteins still formed s odium dodecyl sulfate-stable oligomers but were unable to bind to cells. Ou r data indicate that the C terminus of C2II mediates binding of the protein to cells and that the 7 C-terminal amino acids are structurally important for receptor binding.