Abstract
Fragments of Clostridium botulinum neurotoxin A (BoNT/A) gene (botA) were expressed in Listeria monocytogenes ATCC10527 to produce the L-chain of the toxin in a soluble form. A shuttle vector pAT19 (EmR) was used to make plasmid pAT-RL containing a botA gene fragment placed under C. botulinum ntnH-gene promoter control. The plasmid also contained a C. botulinum botR/A gene, a positive transcriptional regulator of botA. The cytoplasmic fraction of the L. monocytogenes (pAT-RL) strain was found to contain up to 3 mg/L of a soluble protein of expected size and immunologically positive towards BoNT antibodies. This is the first evidence of heterologous botA gene expression producing a soluble safe derivative of botulinum neurotoxin A needed as a molecular tool for exploratory research in neurosciences as well as a basis for raising protective immunity in humans.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Antibodies, Bacterial / blood
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Botulinum Toxins, Type A / biosynthesis*
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Botulinum Toxins, Type A / chemistry
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Botulinum Toxins, Type A / genetics
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Clostridium botulinum / chemistry
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Clostridium botulinum / genetics*
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DNA, Bacterial / chemistry
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DNA, Bacterial / genetics
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DNA, Bacterial / isolation & purification
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Electrophoresis, Polyacrylamide Gel
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Escherichia coli / chemistry
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Escherichia coli / genetics
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Listeria monocytogenes / chemistry
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Listeria monocytogenes / genetics*
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Listeria monocytogenes / metabolism
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Peptide Fragments / biosynthesis
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Peptide Fragments / chemistry
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Peptide Fragments / genetics
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Plasmids / chemistry
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Plasmids / genetics
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Polymerase Chain Reaction
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Rabbits
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Recombinant Proteins / chemistry
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Recombinant Proteins / genetics
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Toxoids / biosynthesis
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Transformation, Bacterial / genetics
Substances
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Antibodies, Bacterial
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DNA, Bacterial
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Peptide Fragments
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Recombinant Proteins
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Toxoids
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Botulinum Toxins, Type A