Abstract
The Lactococcus lactis bacterium found in different natural environments is traditionally associated with the fermented food industry. But recently, its applications have been spreading to the pharmaceutical industry, which has exploited its probiotic characteristics and is moving towards its use as cell factories for the production of added-value recombinant proteins and plasmid DNA (pDNA) for DNA vaccination, as a safer and industrially profitable alternative to the traditional Escherichia coli host. Additionally, due to its food-grade and generally recognized safe status, there have been an increasing number of studies about its use in live mucosal vaccination. In this review, we critically systematize the plasmid replicons available for the production of pharmaceutical-grade pDNA and recombinant proteins by L. lactis. A plasmid vector is an easily customized component when the goal is to engineer bacteria in order to produce a heterologous compound in industrially significant amounts, as an alternative to genomic DNA modifications. The additional burden to the cell depends on plasmid copy number and on the expression level, targeting location and type of protein expressed. For live mucosal vaccination applications, besides the presence of the necessary regulatory sequences, it is imperative that cells produce the antigen of interest in sufficient yields. The cell wall anchored antigens had shown more promising results in live mucosal vaccination studies, when compared with intracellular or secreted antigens. On the other side, engineering L. lactis to express membrane proteins, especially if they have a eukaryotic background, increases the overall cellular burden. The different alternative replicons for live mucosal vaccination, using L. lactis as the DNA vaccine carrier or the antigen producer, are critically reviewed, as a starting platform to choose or engineer the best vector for each application.
Highlights
Introduction published maps and institutional affilLactic Acid Bacteria (LAB) constitute a heterogeneous group of Gram-positive bacteria that share the ability to produce lactic acid as a major product from the fermentation of sugars
The recombinant bacterial strain was orally administered to mice and significant levels of IL-1 receptor antagonist (IL-1Ra) were detected in the mice colon, where it inhibited the IL-1 signaling and alleviated the acute colitis symptoms. These results proves that recombinant L. lactis are able to reach the colon alive and secrete IL-1Ra in situ
The data analyzed in the present review give an important insight about the more appropriate L. lactis vectors to choose or how to engineer them when the goal is to produce high amounts of plasmid DNA (pDNA) and/or recombinant protein, with high quality standards for pharmaceutical or industrial applications
Summary
The efficient protein secretion system of L. lactis confers to this species a major advantage as a protein production platform [7], unlike E. coli for which the most commonly used production strategies are intracellular (periplasm or cytoplasm), leading to expensive downstream purification processes. Concerning protein production, L. lactis is an attractive alternative to the competitor Gram-positive bacterium Bacillus subtilis, the main drawback of which is the degradation of the secreted recombinant proteins by its complex extracellular proteolytic system [12]. Gram-positive bacteria, such as Mycobacterium or Streptococcus, have been studied as hosts for heterologous protein production [14], but the L. lactis food-grade and GRAS status turn them into a valuable alternative. The majority of the plasmids had a pSH71 replicon and further information about it will be provided latter
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