A dileucine in the protease of botulinum toxin a underlies its long-lived neuroparalysis
Wang, Jiafu Zurawski, Tomas H. Meng, Jianghui Lawrence, Gary Olango, Weredeselam M. Finn, David P. Wheeler, Larry Dolly, J. Oliver
Wang, Jiafu
Zurawski, Tomas H.
Meng, Jianghui
Lawrence, Gary
Olango, Weredeselam M.
Finn, David P.
Wheeler, Larry
Dolly, J. Oliver
Identifiers
http://hdl.handle.net/10379/14366
https://doi.org/10.13025/28054
https://doi.org/10.13025/28054
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Publication Date
2010-12-07
Type
Article
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Wang, Jiafu; Zurawski, Tomas H. Meng, Jianghui; Lawrence, Gary; Olango, Weredeselam M.; Finn, David P.; Wheeler, Larry; Dolly, J. Oliver (2010). A dileucine in the protease of botulinum toxin a underlies its long-lived neuroparalysis. Journal of Biological Chemistry 286 (8), 6375-6385
Abstract
Blockade of neurotransmitter release by botulinum neurotoxin type A (BoNT(A)) underlies the severe neuroparalytic symptoms of human botulism, which can last a few years. The structural basis for this remarkable persistence remains unclear. Herein, recombinant BoNT(A) was found to match the neurotoxicity of that from Clostridium botulinum, producing persistent cleavage of synaptosomal-associated protein of 25 kDa (SNAP-25) and neuromuscular paralysis. When two leucines near the C terminus of the protease light chain of A (LC(A)) were mutated, its inhibition of exocytosis was followed by fast recovery of intact SNAP-25 in cerebellar neurons and neuromuscular transmission in vivo. Deletion of 6-7 N terminus residues diminished BoNT(A) activity but did not alter the longevity of its SNAP-25 cleavage and neuromuscular paralysis. Furthermore, genetically fusing LC(E) to a BoNT(A) enzymically inactive mutant (BoTIM(A)) yielded a novel LC(E)-BoTIM(A) protein that targets neurons, and the BoTIM(A) moiety also delivers and stabilizes the inhibitory LC(E), giving a potent and persistent cleavage of SNAP-25 with associated neuromuscular paralysis. Moreover, its neurotropism was extended to sensory neurons normally insensitive to BoNT(E). LC(E)-BoTIM(A)(AA) with the above-identified dileucine mutated gave transient neuromuscular paralysis similar to BoNT(E), reaffirming that these residues are critical for the persistent action of LC(E)-BoTIM(A) as well as BoNT(A). LC(E)-BoTIM(A) inhibited release of calcitonin gene-related peptide from sensory neurons mediated by transient receptor potential vanilloid type 1 and attenuated capsaicin-evoked nociceptive behavior in rats, following intraplantar injection. Thus, a long acting, versatile composite toxin has been developed with therapeutic potential for pain and conditions caused by overactive cholinergic nerves.
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Publisher
American Society for Biochemistry & Molecular Biology (ASBMB)
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Attribution-NonCommercial-NoDerivs 3.0 Ireland