Neuronal entry and high neurotoxicity of botulinum neurotoxin A require its N-terminal binding sub-domain
| Autor: | J. Oliver Dolly, Jianghui Meng, Marc Nugent, Minhong Tang, Jiafu Wang |
|---|---|
| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
Sensory Receptor Cells media_common.quotation_subject Primary Cell Culture Mutant Synaptic vesicle Article 03 medical and health sciences 0302 clinical medicine Protein Domains medicine Animals Humans Paralysis Botulinum Toxins Type A Internalization media_common Neurons Membrane Glycoproteins Multidisciplinary Chemistry Cell Membrane Neurotoxicity Wild type medicine.disease Acetylcholine Recombinant Proteins Rats 3. Good health Cell biology 030104 developmental biology Membrane protein Cell culture Mutant Proteins Synaptic Vesicles 030217 neurology & neurosurgery Protein Binding medicine.drug |
| Zdroj: | Scientific Reports |
| ISSN: | 2045-2322 |
| DOI: | 10.1038/srep44474 |
| Popis: | Botulinum neurotoxins (BoNTs) are the most toxic proteins known, due to inhibiting the neuronal release of acetylcholine and causing flaccid paralysis. Most BoNT serotypes target neurons by binding to synaptic vesicle proteins and gangliosides via a C-terminal binding sub-domain (HCC). However, the role of their conserved N-terminal sub-domain (HCN) has not been established. Herein, we created a mutant form of recombinant BoNT/A lacking HCN (rAΔHCN) and showed that the lethality of this mutant is reduced 3.3 × 104-fold compared to wild-type BoNT/A. Accordingly, low concentrations of rAΔHCN failed to bind either synaptic vesicle protein 2C or neurons, unlike the high-affinity neuronal binding obtained with 125I-BoNT/A (Kd = 0.46 nM). At a higher concentration, rAΔHCN did bind to cultured sensory neurons and cluster on the surface, even after 24 h exposure. In contrast, BoNT/A became internalised and its light chain appeared associated with the plasmalemma, and partially co-localised with vesicle-associated membrane protein 2 in some vesicular compartments. We further found that a point mutation (W985L) within HCN reduced the toxicity over 10-fold, while this mutant maintained the same level of binding to neurons as wild type BoNT/A, suggesting that HCN makes additional contributions to productive internalization/translocation steps beyond binding to neurons. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|