Effective expression of fragments of a botulinum neurotoxin type A gene, coding for the L-chain and H-chain in E. coli, with formation of products causing protective immunity to administration of the toxin

Native Clostridium botulinum gene coding for type A neurotoxin has been used to construct recombinant derivatives coding separately for L and H polypeptide chains of the toxin. The gene derivatives have been cloned into an expression vector pET28b in E. coli BL21 (DE3) cells. The recombinant L and H...

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Bibliographic Details
Published in:	Molekulârnaâ genetika, mikrobiologiâ i virusologiâ no. 4; p. 3
Main Authors:	Vertiev, Iu V, Zdanovskiĭ, A G, Borinskaia, S A, Martin, T, Gening, E L, Iankovskiĭ, N K
Format:	Journal Article
Language:	Russian
Published:	Russia (Federation) 2000
Subjects:	Base Sequence Botulinum Toxins - administration & dosage Botulinum Toxins - chemistry Botulinum Toxins - genetics Botulinum Toxins - immunology Cloning, Molecular DNA, Bacterial Escherichia coli - genetics Recombinant Proteins - administration & dosage Recombinant Proteins - chemistry Recombinant Proteins - genetics Recombinant Proteins - immunology
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Summary:	Native Clostridium botulinum gene coding for type A neurotoxin has been used to construct recombinant derivatives coding separately for L and H polypeptide chains of the toxin. The gene derivatives have been cloned into an expression vector pET28b in E. coli BL21 (DE3) cells. The recombinant L and H proteins seem to be the major individual proteins after IPTG induction of the recombinant cells. Each of the proteins has been accumulated only in inclusion bodies. The recombinant L chain (but not H chain) has been successfully resolubilized. Each of the proteins contains six His residues on the N terminus which allows purification on Ni-agarose columns with high yield. No toxic effect has been observed for both L and H chains after injection of 10 micrograms of recombinant preparations purified from inclusion bodies. Moreover, the injection resulted in an increase in the titer of specific antibodies which protected mice from 1 DLM of type A native botulinum neurotoxin. Hence, the recombinant neurotoxin protein derivatives which are present in E. coli inclusion bodies can be a source of material for producing diagnostic and therapeutic sera against type A botulinum neurotoxin.
ISSN:	0208-0613