THE N-TERMINAL PART OF THE ENZYME COMPONENT (C2I) OF THE BINARY CLOSTRIDIUM-BOTULINUM C2 TOXIN INTERACTS WITH THE BINDING-COMPONENT C2II AND FUNCTIONS AS A CARRIER SYSTEM FOR A RHO ADP-RIBOSYLATING C3-LIKE FUSION TOXIN

The binary actin-ADP-ribosylating Clostridium botulinum C2 toxin consi sts of the enzyme component C2I and the binding component C2II, which are separate proteins. The active component C2I enters cells through C 2II by receptor-mediated endocytosis and membrane translocation. The N -terminal part of C2I (C2IN), which consists of 225 amino acid residue s but lacks ADP-ribosyltransferase activity, was identified as the C2I I contact site. A fusion protein (C2IN-C3) of C2IN and the full-length C3-like ADP-ribosyltransferase from Clostridium limosum was construct ed. The fusion protein C2IN-C3 ADP-ribosylated Rho but not actin in CH O cell lysates. Together with C2II, C2IN-C3 induced complete rounding up of CHO and HeLa cells after incubation for 3 h. No cell rounding wa s observed without C2II or with the original C3-like transferase from C. limosum. The data indicate that the N-terminal 225 amino acid resid ues of C2I are sufficient to cause the cellular uptake of C. limosum t ransferase via the binding component of C2II, thereby increasing the c ytotoxicity of the C3-like exoenzyme several hundred fold.