Dataset Information

Comparative genome-based identification of a cell wall-anchored protein from Lactobacillus plantarum increases adhesion of Lactococcus lactis to human epithelial cells.

ABSTRACT: Adhesion to host cells is considered important for Lactobacillus plantarum as well as other lactic acid bacteria (LAB) to persist in human gut and thus exert probiotic effects. Here, we sequenced the genome of Lt. plantarum strain NL42 originating from a traditional Chinese dairy product, performed comparative genomic analysis and characterized a novel adhesion factor. The genome of NL42 was highly divergent from its closest neighbors, especially in six large genomic regions. NL42 harbors a total of 42 genes encoding adhesion-associated proteins; among them, cwaA encodes a protein containing multiple domains, including five cell wall surface anchor repeat domains and an LPxTG-like cell wall anchor motif. Expression of cwaA in Lactococcus lactis significantly increased its autoaggregation and hydrophobicity, and conferred the new ability to adhere to human colonic epithelial HT-29 cells by targeting cellular surface proteins, and not carbohydrate moieties, for CwaA adhesion. In addition, the recombinant Lc. lactis inhibited adhesion of Staphylococcus aureus and Escherichia coli to HT-29 cells, mainly by exclusion. We conclude that CwaA is a novel adhesion factor in Lt. plantarum and a potential candidate for improving the adhesion ability of probiotics or other bacteria of interest.

SUBMITTER: Zhang B

PROVIDER: S-EPMC4572922 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Comparative genome-based identification of a cell wall-anchored protein from Lactobacillus plantarum increases adhesion of Lactococcus lactis to human epithelial cells.

Zhang Bo B Zuo Fanglei F Yu Rui R Zeng Zhu Z Ma Huiqin H Chen Shangwu S

Scientific reports 20150915

Adhesion to host cells is considered important for Lactobacillus plantarum as well as other lactic acid bacteria (LAB) to persist in human gut and thus exert probiotic effects. Here, we sequenced the genome of Lt. plantarum strain NL42 originating from a traditional Chinese dairy product, performed comparative genomic analysis and characterized a novel adhesion factor. The genome of NL42 was highly divergent from its closest neighbors, especially in six large genomic regions. NL42 harbors a tota ...[more]

PMID: 26370773

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Project description:Campylobacter jejuni is the most frequent cause of human food-borne gastroenteritis and chicken meat is the main source of infection. Recent studies showed that broiler chicken immunization against Campylobacter should be the most efficient way to lower the number of human infections by this pathogen. Induction of the mucosal immune system after oral antigen administration should provide protective immunity to chickens. In this work we tested the usefulness of Lactococcus lactis, the most extensively studied lactic acid bacterium, as a delivery vector for Campylobacter antigens. First we constructed hybrid protein - CjaA antigen presenting CjaD peptide epitopes on its surface. We showed that specific rabbit anti-rCjaAD serum reacted strongly with both CjaA and CjaD produced by a wild type C. jejuni strain. Next, rCjaAD and CjaA were fused to the C-terminus of the L. lactis YndF containing the LPTXG motif. The genes expressing these proteins were transcribed under control of the L. lactis Usp45 promoter and their products contain the Usp45 signal sequences. This strategy ensures a cell surface location of both analyzed proteins, which was confirmed by immunofluorescence assay. In order to evaluate the impact of antigen location on vaccine prototype efficacy, a L. lactis strain producing cytoplasm-located rCjaAD was also generated. Animal experiments showed a decrease of Campylobacter cecal load in vaccinated birds as compared with the control group and showed that the L. lactis harboring the surface-exposed rCjaAD antigen afforded greater protection than the L. lactis producing cytoplasm-located rCjaAD. To the best of our knowledge, this is the first attempt to employ Lactic Acid Bacteria (LAB) strains as a mucosal delivery vehicle for chicken immunization. Although the observed reduction of chicken colonization by Campylobacter resulting from vaccination was rather moderate, the experiments showed that LAB strains can be considered as an alternative vector to deliver heterologous antigens to the bird immune system. Additionally, the analysis of the structure and immunogenicity of the generated rCjaAD hybrid protein showed that the CjaA antigen can be considered as a starting point to construct multiepitope anti-Campylobacter vaccines.

Project description:Large-scale mass spectrometry-based peptidomics for bioactive-peptide discovery is relatively unexplored because of challenges in intracellular peptide extraction and small-peptide identification. Here, we present an analytical pipeline for large-scale intracellular peptidomics of Lactococcus lactis It entails an optimized sample preparation protocol for L. lactis, used as an "enzyme complex" to digest β-casein, an extraction method for its intracellular peptidome, and a peptidomics data analysis and visualization procedure. In addition, we proofread the publicly available bioactive-peptide databases and obtained an optimized database of bioactive peptides derivable from bovine β-casein. We used the pipeline to examine cultures of L. lactis MG1363 and a set of 6 isogenic multiple peptidase mutants incubated with β-casein. We observed a clearly strain-dependent accumulation of peptides with several bioactivities, such as angiotensin-converting enzyme (ACE)-inhibitory, dipeptidyl peptidase 4 (DPP-IV)-inhibitory, and immunoregulatory functions. The results suggest that both the number of different bioactive peptides and the bioactivity diversity can be increased by editing the proteolytic system of L. lactis This comprehensive pipeline offers a model for discovery of bioactive peptides in combination with other proteins and might be applicable to other bacteria.IMPORTANCE Lactic acid bacteria (LAB) are very important for the production of safe and healthy human and animal fermented foods and feed and, increasingly more, in the functional food industry. The intracellular peptidomes of LAB are promising reservoirs of bioactive peptides. We show here that targeted genetic engineering of the peptide degradation pathway allows steering the composition of the peptide pool of the LAB Lactococcus lactis and production of peptides with interesting bioactivities. Our work could be used as a guideline for modifying proteolytic systems in other LAB to further explore their potential as cell peptide factories.

Dataset Information

Comparative genome-based identification of a cell wall-anchored protein from Lactobacillus plantarum increases adhesion of Lactococcus lactis to human epithelial cells.

Publications

Comparative genome-based identification of a cell wall-anchored protein from Lactobacillus plantarum increases adhesion of Lactococcus lactis to human epithelial cells.

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