Characterization, cloning, curing, and distribution in lactic acid bacteria of pLP1, a plasmid from Lactobacillus plantarum CCM 1904 and its use in shuttle vector construction

Bringel, Françoise; Frey, Lucie; Hubert, J.

doi:10.1016/0147-619x(89)90002-4

Cited by 48 publications

(16 citation statements)

References 26 publications

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“…The protocol for plasmid extraction from L. casei and L. plantarum was similar to that used for L. sakei. Plasmids were established by electroporation as described for L. lactis and S. thermophilus (20), L. reuteri (4), and L. casei and L. sakei (2). For L. plantarum, the following protocol was used (R. Jimenez-Diaz, personal communication).…”

Section: Methodsmentioning

confidence: 99%

Design of a Protein-Targeting System for Lactic Acid Bacteria

et al. 2001

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We designed an expression and export system that enabled the targeting of a reporter protein (the staphylococcal nuclease Nuc) to specific locations in Lactococcus lactis cells, i.e., cytoplasm, cell wall, or medium. Optimization of protein secretion and of protein cell wall anchoring was performed with L. lactis cells by modifying the signals located at the N and C termini, respectively, of the reporter protein. Efficient translocation of precursor (ϳ95%) is obtained using the signal peptide from the lactococcal Usp45 protein and provided that the mature protein is fused to overall anionic amino acids at its N terminus; those residues prevented interactions of Nuc with the cell envelope. Nuc could be covalently anchored to the peptidoglycan by using the cell wall anchor motif of the Streptococcus pyogenes M6 protein. However, the anchoring step proved to not be totally efficient in L. lactis, as considerable amounts of protein remained membrane associated. Our results may suggest that the defect is due to limiting sortase in the cell. The optimized expression and export vectors also allowed secretion and cell wall anchoring of Nuc in food-fermenting and commensal strains of Lactobacillus. In all strains tested, both secreted and cell wall-anchored Nuc was enzymatically active, suggesting proper enzyme folding in the different locations. These results provide the first report of a targeting system in lactic acid bacteria in which the final location of a protein is controlled and biological activity is maintained.

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Section: Methodsmentioning

confidence: 99%

Design of a Protein-Targeting System for Lactic Acid Bacteria

et al. 2001

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“…Both RC and theta types of replicons have been reported in lactobacilli. In L. sakei, RC plasmids closely related to those of Lactobacillus curvatus (31) and Lactobacillus plantarum (9) have been identified, and the partial sequence of a potential theta-replicating plasmid is known, namely, pSAK1 (GenBank Z50862), belonging to the pLS32 family (49). In this paper, we present the analysis of the complete sequence of a new L. sakei plasmid.…”

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confidence: 99%

Characterization of a Theta-Type Plasmid from Lactobacillus sakei : a Potential Basis for Low-Copy-Number Vectors in Lactobacilli

Alpert¹,

Coq²,

Malleret³

et al. 2003

Appl Environ Microbiol

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The complete nucleotide sequence of the 13-kb plasmid pRV500, isolated from Lactobacillus sakei RV332, was determined. Sequence analysis enabled the identification of genes coding for a putative type I restrictionmodification system, two genes coding for putative recombinases of the integrase family, and a region likely involved in replication. The structural features of this region, comprising a putative ori segment containing 11-and 22-bp repeats and a repA gene coding for a putative initiator protein, indicated that pRV500 belongs to the pUCL287 subfamily of theta-type replicons. A 3.7-kb fragment encompassing this region was fused to an Escherichia coli replicon to produce the shuttle vector pRV566 and was observed to be functional in L. sakei for plasmid replication. The L. sakei replicon alone could not support replication in E. coli. Plasmid pRV500 and its derivative pRV566 were determined to be at very low copy numbers in L. sakei. pRV566 was maintained at a reasonable rate over 20 generations in several lactobacilli, such as Lactobacillus curvatus, Lactobacillus casei, and Lactobacillus plantarum, in addition to L. sakei, making it an interesting basis for developing vectors. Sequence relationships with other plasmids are described and discussed.Lactobacilli belong to the lactic acid bacteria (LAB) group. The genus comprises several species important for the food industry, among which is Lactobacillus sakei, one of the most important bacterial species involved in meat fermentation and preservation. L. sakei belongs to the dominant microflora naturally occurring on vacuum-packed meat and is also used in Western Europe as starter cultures for fermented products (especially sausage) because of its ability to rapidly ferment carbohydrates in the meat environment (11,26).Most lactobacilli harbor one or more plasmids, the size of which can vary from 1.2 to 150 kb (for a review, see reference 50). Different functions have already been found on these plasmids, among which are genes for lactose metabolism, bacteriocin synthesis, exopolysaccharide production, or DNA restriction-modification (R-M). With the development of molecular biology and functional genomics of lactobacilli, interest is growing for the characterization of replicons themselves as potential useful (food-grade) vectors. Two modes of DNA replication are used by circular bacterial plasmids, namely, rolling circle (RC) and theta. RC plasmids have been assumed to be the most widespread in gram-positive bacteria. However, recent years have seen the characterization of a large number of theta-replicating plasmids from gram-positive, and especially from LAB, hosts. Several mechanisms are known for the initiation of theta replication. Theta replicons can be classified according to their dependence on three key components. These are plasmid-encoded initiator Rep proteins necessary for strand opening and/or interaction with other components of the initiation complex, origins of replication (generically termed ori) with specific DNA structural organization...

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“…This characteristic of Lactobacillus supplies a large reservoir of plasmids which can be exploited for the construction of the third class of Lactobacillus vector, and for diverse recombinant applications. Since repA-based plasmid replicons replicate via the theta mode of replication, which confers stability, derivatives of this replicons type have been widely used (86)(87)(88)(99)(100)(101) (102). L. salivarius UCC118 harbours 3 plasmids (36), a megaplasmid pMP118 and two endogenous plasmids pSF118-44 and pSF118-20 which putatively replicate via a theta replication mechanism.…”

Section: Available Plasmids Replicons and Construction Of Gene Cloninmentioning

confidence: 99%

Genetic tools for investigating the biology of commensal lactobacilli

Fang

O’Toole²

2009

Front Biosci

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Lactobacilli belong to the genus Lactobacillus, the largest genus among the lactic acid bacteria (LAB). They are abundant in plant material and food resources, or they may inhabit niches in or on the bodies of humans and animals, as commensals. Lactobacilli of food origin are commercially important in the production of dairy products, fermented meats, vegetables, and sourdough, and many of their properties have been well studied. Commensal lactobacilli are good candidates for development as probiotics. In recent years, the general biology and host interaction mechanisms of commensal lactobacilli have attracted great interest. Although the metabolic pathways, predicted gene functions, and some phenotypic traits, of commensal lactobacilli can be inferred or deduced to an extent by the growing number of Lactobacillus genome sequencing project, various genetic tools are still required to confirm their phenotypic properties and biological traits. The current state of the art with respect to the available complement of genetic tools including genomic resources, and more traditional approaches to investigate the biology of commensal lactobacilli, will now be reviewed

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Characterization, cloning, curing, and distribution in lactic acid bacteria of pLP1, a plasmid from Lactobacillus plantarum CCM 1904 and its use in shuttle vector construction

Cited by 48 publications

References 26 publications

Design of a Protein-Targeting System for Lactic Acid Bacteria

Design of a Protein-Targeting System for Lactic Acid Bacteria

Characterization of a Theta-Type Plasmid from Lactobacillus sakei : a Potential Basis for Low-Copy-Number Vectors in Lactobacilli

Genetic tools for investigating the biology of commensal lactobacilli

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