Translocation domain of botulinum neurotoxin A subtype 2 potently induces entry into neuronal cells.

Botulinum neurotoxin (BoNT) is the causative agent of botulism in humans and animals. Only BoNT serotype A subtype 1 (BoNT/A1) is clinically used because of its high potency and long duration of action. BoNT/A1 and BoNT/A subtype 2 (BoNT/A2) have high degree of amino acid sequence similarity in light chain (LC) (96%), whereas their N-and C-terminal heavy chain (HN and HC ) differ by 13%. LC acts as a zinc-dependent endopeptidase, HN as the translocation domain, and HC as the receptor-binding domain. BoNT/A2 and BoNT/A1 had similar potency in mouse bioassay, but BoNT/A2 entered faster and more efficiently into neuronal cells. To identify domains responsible for these characteristics, HN of BoNT/A1 and BoNT/A2 was exchanged to construct chimeric BoNT/A121 and BoNT/A212. After expression in E. coli, chimeric and wild-type BoNT/As were purified as single-chain proteins and activated by conversion to disulfide-linked dichains. Toxicities of recombinant wild-type and chimeric BoNT/As were similar, but it dropped to 60% compared with the values of native BoNT/As. The relative orders of SNAP-25 cleavage activity in neuronal cells and toxicity differed. BoNT/A121 and recombinant BoNT/A2 have similar SNAP-25 cleavage activity. BoNT/A2 HN is possibly responsible for the higher potency of BoNT/A2 than BoNT/A1. This article is protected by copyright. All rights reserved.