Endopeptidase Activities of Botulinum Neurotoxin Type B Complex, Holotoxin, and Light Chain
- 1 October 2010
- journal article
- research article
- Published by American Society for Microbiology in Applied and Environmental Microbiology
- Vol. 76 (19), 6658-6663
- https://doi.org/10.1128/aem.00731-10
Abstract
Botulinum neurotoxin (BoNT) serotype B (BoNT/B) is one of the serotypes of BoNT that causes deadly human botulism, though it is used clinically for treatment of many neuromuscular diseases. BoNT/B is produced by Clostridium botulinum , and it is secreted along with a group of neurotoxin-associated proteins (NAPs) in the form of a BoNT/B complex. The complex dissociates into a 150-kDa holotoxin and NAPs at alkaline pHs. The 150-kDa BoNT/B holotoxin can be nicked to produce a 50-kDa domain referred to as the light chain (LC) and a 100-kDa heavy chain, with the former possessing a unique endopeptidase activity. The two chains remain linked through a disulfide bond that can be reduced to separate the two chains. The endopeptidase activity is present in all three forms of the toxin (complex, purified BoNT/B holotoxin, and separated light chain), which are used by different researchers to develop detection methods and screen for inhibitors. In this research, the endopeptidase activities of the three forms, for the first time, were compared under the same conditions. The results show that enzyme activities of the three forms differ significantly and are largely dependent on nicking and disulfide reduction conditions. Under the conditions used, LC had the highest level of activity, and the complex had the lowest. The activity was enhanced by nicking of BoNT/B holotoxin and was enhanced even more by dithiothreitol (DTT) reduction after nicking. This information is useful for understanding the properties of BoNT endopeptidases and for comparing the efficacies of different inhibitors when they are tested with different forms of BoNT endopeptidase.Keywords
This publication has 15 references indexed in Scilit:
- How do the Botulinum Neurotoxins block neurotransmitter release: from botulism to the molecular mechanism of actionThe Botulinum J., 2008
- Detection of Type A, B, E, and F Clostridium botulinum Neurotoxins in Foods by Using an Amplified Enzyme-Linked Immunosorbent Assay with Digoxigenin-Labeled AntibodiesApplied and Environmental Microbiology, 2006
- Comparison of botulinum toxin serotypes A and B for the treatment of cervical dystoniaNeurology, 2005
- Expression, purification, and characterization of Clostridium botulinum type B light chainProtein Expression and Purification, 2005
- Analysis of the substrate recognition domain determinants of Botulinum Type B toxin using Phage DisplayToxicon, 2005
- Enhancement of the Endopeptidase Activity of Purified Botulinum Neurotoxins A and E by an Isolated Component of the Native Neurotoxin Associated ProteinsBiochemistry, 2004
- Botulinum toxin type B (Myobloc®): pharmacology and biochemistryClinics in Dermatology, 2004
- Fluorigenic Substrates for the Protease Activities of Botulinum Neurotoxins, Serotypes A, B, and FApplied and Environmental Microbiology, 2003
- Efficacy of a novel metalloprotease inhibitor on botulinum neurotoxin B activityFEBS Letters, 1998
- Peptide Substrate Specificity and Properties of the zinc‐endopeptidase Activity of Botulinum Type B NeurotoxinJBIC Journal of Biological Inorganic Chemistry, 1994