Nisin-controlled expression of Norwalk virus VP60 protein in

Abstract

A food-grade strain with nisRK stably integrated into the genome, was constructed in order to implement the nisin-controlled expression system (NICE) in Lactobacillus casei ATCC393. Expression of beta-glucuronidase (gus) reporter gene was employed to optimize the system, which has been successfully used to produce the main antigenic protein from Norwalk virus, opening new perspectives for producing edible vaccines.

Generation of food-grade recombinant Lactobacillus casei delivering Myxococcus xanthus prolyl endopeptidase

“…Thus, L. casei BL23 (Mazé et al 2010) was electroporated with both, pIPLA1503 and pIPLA1506 plasmids. The resulting strains were transformed with the replicative plasmid pEM94 (Martin et al 2004) which contains the gene encoding the β -recombinase, with the aim of removing by site-specific recombination, the non-food grade DNA located between two six sites. To eliminate pEM94, which carries a temperature sensitive origin of replication, the strains were cultured at 37° C. The resulting strains were designated as L. casei IPLA1503 and L. casei IPLA1506, respectively.…”

“…To achieve this goal, we had to integrate the pep gene stably into the chromosome, keep the food-grade status of the strain, produce the enzyme in sufficient quantity constitutively, and produce the enzyme both in the cytoplasm and in the extracellular media. The two first challenges were overcome via the food-grade integrative system of Martin and coworkers (Martin et al 2000; Martin et al 2004; Martin et al 2011). This system was used to deliver a codon-optimized gene encoding Mx PEP under the control of the constitutive apf promoter of Lactobacillus crispatus (Martin et al 2011), and using the apf signal peptide, it was possible to secrete the 76.8 kDa protein outside the cell.…”