Recombinant Listeria strains producing the nontoxic L-chain of botulinum neurotoxin A in a soluble form

Fragments of Clostridium botulinum neurotoxin A (BoNT/A) gene (botA) were e xpressed in Listeria monocytogenes ATCC10527 to produce the L-chain of the toxin in a soluble form. A shuttle vector pAT19 (Em(R)) was used to make pl asmid pAT-RL containing a botA gene fragment placed under C. botulinum ntnH -gene promoter control. The plasmid also contained a C. botulinum botR/A ge ne, 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 o f a soluble protein of expected size and immunologically positive towards B oNT antibodies. This is the first evidence of heterologous botA gene expres sion 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. (C) 2001 Editions scientif iques et medicales Elsevier SAS.