ABSTRACT
Botulinum neurotoxin (BoNT) binds to presynaptic neuronal cells and blocks neurotransmitter release. The carboxyl-terminal half of the heavy chain (H(C)) of the neurotoxin recognizes its specific receptor on the plasma membrane. We have previously demonstrated that BoNT/C binds to gangliosides GD1b and GT1b under physiological conditions, while BoNT/D interacts with phosphatidylethanolamine (PE). Here we report that the recognition sites for gangliosides and PE are present in the carboxyl-terminal domain of H(C). Chimeric mutants and site-directed mutants of BoNT/C-H(C) and BoNT/D-H(C) were generated and their binding activities evaluated. The chimeric H(C) that consisted of the amino-terminal half of BoNT/D-H(C) and the carboxyl-terminal half of BoNT/C-H(C) possessed activity similar to the authentic BoNT/C-H(C), suggesting that the carboxyl-terminal region of H(C) is involved in the receptor recognition of BoNT/C. Moreover, analysis using site-directed mutants indicated that the peptide motif W(1257)Ycdots, three dots, centeredG(1270)cdots, three dots, centeredH(1282) plays an important role in the interaction between BoNT/C and gangliosides. In contrast, we revealed that two lysine residues of BoNT/D-H(C) are involved in the formation of the critical binding site for receptor binding.
