Expression and purification of catalytically active, non-toxic endopeptidase derivatives of Clostridium botulinum toxin type A.

摘要: Abstract Clostridium botulinum neurotoxin type A is a potently toxic protein of 150 kDa with specific endopeptidase activity for the SNARE SNAP-25. Proteolytic cleavage BoNT/A trypsin leads to removal C-terminal domain responsible neuronal cell binding. Removal this result in catalytically active, non-cell-binding derivative termed LHN/A. We have developed purification scheme prepare LHN/A essentially free contaminating BoNT/A. prepared by retains full enzymatic activity, stable solution, and low toxicity as demonstrated mouse assay. In addition, has minimal effect on release neurotransmitter from primary culture model. Both bioassay vitro assay suggest present at less than 1 106 molecules This represents significant improvement previously reported figures LHN/A, also light chain domain, purified To complement preparation holotoxin, DNA encoding been introduced into Escherichia coli facilitate expression recombinant product. Expression parameters enable isolation soluble, profile enhanced that The recombinant-derived material used antisera neutralise challenge. production BoNT/A-free two different methods possibilities exploitation are discussed.