A role for the interchain disulfide or its participating thiols in the internalization of botulinum neurotoxin A revealed by a toxin derivative that binds to ecto-acceptors and inhibits transmitter release intracellularly - PubMed (original) (raw)

. 1993 Oct 5;268(28):20838-44.

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A role for the interchain disulfide or its participating thiols in the internalization of botulinum neurotoxin A revealed by a toxin derivative that binds to ecto-acceptors and inhibits transmitter release intracellularly

A de Paiva et al. J Biol Chem. 1993.

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Abstract

Botulinum neurotoxin type A consists of a disulfide-linked light and heavy chain, with an intradisulfide present within the C-terminal half of the latter. The functional consequences of reducing these bonds and alkylating the thiols were investigated. Modification of free cysteine residues had no effect on the toxicity in mouse bioassays or on acetylcholine release in the mouse nerve-diaphragm and the buccal ganglion of Aplysia californica. However, reduction of the toxin prior to alkylation drastically decreased neuroparalytic potency; yet, this derivative inhibited transmitter release if injected directly into a presynaptic neuron in the Aplysia ganglion or added to bovine permeabilized adrenal chromaffin cells. Its antagonism of the action of botulinum neurotoxin A at mammalian motor nerve endings and Aplysia neurons indicates retention of the ability to bind to the toxin's productive ecto-acceptors. Thus, the abolition of the toxicity of extracellularly applied botulinum neurotoxin A by the cleavage of both disulfides, and the alkylation of the half-cystines involved, results from ineffective uptake. Modified forms of the isolated chains of botulinum neurotoxin A were utilized to determine which of the disulfides were necessary for internalization. Alkylation of the cysteines in the light and heavy chains, including those involved in the interchain bond but excluding those of the intact disulfide in the heavy chain, revealed that the intermolecular bond must be present, or the thiols concerned unmodified, for botulinum neurotoxin A to undergo membrane translocation into Aplysia neurons.

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