Novel Treatments for Botulism: Development of Antagonists for Identified Steps in the Action of Botulinum Neurotoxins
Abstract
With the aim of identifying domains in botulinum neurotoxin (BoNT) responsible for blockade of transmitter release, a prerequisite for developing treatments for botulism, the heavy (HC) and light (LC) chains of types A and B were purified to homogeneity, renatured, or reconstituted to give a di-chain toxic species; controlled proteolysis of BoNT A followed by chromatographic procedures yielded pure fragments (H2L, intact toxin minus the C-terminal half of HC (H1); H2, N-terminal portion of HC). These were tested, alone or in combination, for inhibitory effects on neurally-evoked transmitter release from mouse nerve diaphragm and Aplysia ganglionic neurons. The latter allowed extra- or intra-neuronal administration of toxin samples, with quantitation of quantal release by voltage-clamp analysis of pairs of pre- and post-synaptic cells. Examination of an intracellular action of BoNT was, also, accomplished by digitonin-permeabilisation of cultured PC-12 cells, with retention of the exocytosis process. For monitoring the toxins's molecular action, nerve terminals or synaptic vesicles were isolated from brain. Keywords: Botulism; Botulinum neurotoxin; Botulinum neurotoxin acceptors; Internalisation; Antibodies; ADP-ribosylation; Exocytosis; Neuromuscular junction; Acetylcholine; Noradrena-line; G-proteins.
Document Details
- Document Type
- Technical Report
- Publication Date
- Nov 07, 1989
- Accession Number
- ADA215346
Entities
People
- J. O. Dolly
Organizations
- Imperial College London