[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 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.
Vertiev
Iu V
IuV
N. F. Gamaleya Institute of Epidemiology and Microbiology, Russian Academy of Medical Sciences, Moscow, Russia.
Zdanovskiĭ
A G
AG
Borinskaia
S A
SA
Martin
T
T
Gening
E L
EL
Iankovskiĭ
N K
NK
D001483:Base Sequence; D001905:Botulinum Toxins / Q000008:administration & dosage / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D004926:Escherichia coli / Q000235:genetics*; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000737:chemistry / Q000235:genetics / Q000276:immunology
D004269:DNA, Bacterial; D011994:Recombinant Proteins; D001905:Botulinum Toxins