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Purification and Partial Characterization of Bacteriocin Lac-B23, a Novel Bacteriocin Production by Lactobacillus plantarum J23, Isolated From Chinese Traditional Fermented Milk.

ABSTRACT: The exploration and evaluation of bacteriocin-producing lactic acid bacteria (LAB) have been one of the powerful means to food preservation. A total of 300 strains were isolated from Chinese traditional fermented milk products. A bacteriocin-producing LAB, named Lactobacillus plantarum J23, was screened and identified. Bacteriocin Lac-B23 from L. plantarum J23 was purified by 80% ammonium sulfate precipitation, cation-exchange chromatography, and reverse-phase high-performance liquid chromatography. Molecular weight of bacteriocin Lac-B23 was determined to be approximately 6.73 kDa by tricine sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis, and it was confirmed as a novel bacteriocin by liquid chromatography-mass spectrometry. Moreover, bacteriocin Lac-B23 showed thermal stability when heated at below 100°C for 30 min, pH stability between pH 2.0 and 12.0, and sensitivity to trypsin, proteinase K, and proteinase E. The antimicrobial activity of bacteriocin could be enhanced by addition of Fe²⁺, Mn²⁺, and ethyl alcohol, and inhibited by Cu²⁺, K⁺, Ca²⁺, Zn²⁺, Mg²⁺, and sodium chloride. The results suggested bacteriocin Lac-B23 to have potential application prospects in the food industry.

SUBMITTER: Zhang J

PROVIDER: S-EPMC6174205 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Lactobacillus plantarum zrx03 was a bacteriocin-producing strain isolated from infant's feces. The fermentation supernatant produced by this strain could strongly inhibit Escherichia coli JM109 ATCC 67387, Staphylococcus aureus ATCC 25923, and Listeria monocytogenes CICC 21633, in which the diameter of inhibition zone was 12.83 ± 0.62 mm, 15.08 ± 0.31 mm, 6.75 ± 0.20 mm, respectively, compared with lactic acid bacteria N1, N2, M13, M21, M31, and M37. According to amplification of 16S rRNA gene and identification of phylogenetic tree, this strain had a 1,450 bp sequence and 100% identity to the L. plantarum strain. Based on the influence of different protease treatments, such as pepsin, trypsin, papain, and proteinase K on the antimicrobial activity, this antimicrobial substance was considered to be a natural protein. Using bacteriocin produced by this strain as study object of this experiment, it had been extracted from ammonium sulfate precipitation and different organic solvents. The results showed that ethyl acetate was selected as the optimal solution to crude extraction of bacteriocin after comparing ammonium sulfate precipitation method and organic solvent extraction method, such as n-butanol, n-hexane, dichloromethane, trichloromethane, in which the diameter of the inhibition zones was above 28 mm. Results also showed the inhibition spectrum of the obtained bacteriocin had a broad spectrum of inhibition which could inhibit Gram-positive, Gram-negative, yeast. Especially, it could effectively inhibit S. aureus ATCC 25923, Bacillus subtilis CICC 10002, Bacillus anthracis CICC 20443, E. coli JM109 ATCC 67387, and Salmonella CMCC 541, and the zone diameter of inhibition has reached more than 28 mm. Moreover, it had a good thermal stability which antibacterial activity was retained 70.58% after treatment at 121°C for 30 min, and pH-stability was between pH 2.0-9.0. These results suggested bacteriocin produced by L. plantarum zrx03 had potential application prospects in food preservation.

Project description:Bacteriocins from lactic acid bacteria (LAB) are of increasing interest in recent years due to their potential as natural preservatives against food and beverage spoilage microorganisms. In a screening study for LAB, we isolated from olives a strain, Lactobacillus plantarum NI326, with activity against the beverage-spoilage bacterium Alicyclobacillus acidoterrestris Genome sequencing of NI326 enabled the identification of a gene cluster (designated plc) encoding a putative circular bacteriocin and proteins involved in its modification, transport, and immunity. This novel bacteriocin, named plantaricyclin A (PlcA), was grouped into the circular bacteriocin subgroup II due to its high degree of similarity with other gassericin A-like bacteriocins. Purification of PlcA from the supernatant of Lb. plantarum NI326 resulted in an active peptide with a molecular mass of 5,570 Da, corresponding to that predicted from the (processed) PlcA amino acid sequence. The plc gene cluster was cloned and expressed in Lactococcus lactis NZ9000, resulting in the production of an active 5,570-Da bacteriocin in the supernatant. PlcA is believed to be produced as a 91-amino-acid precursor with a 33-amino-acid leader peptide, which is predicted to be removed, followed by joining of the N and C termini via a covalent linkage to form the mature 58-amino-acid circular bacteriocin PlcA. We report the characterization of a circular bacteriocin produced by Lb. plantarum The inhibition displayed against A. acidoterrestris highlights its potential use as a preservative in food and beverages.IMPORTANCE In this work, we describe the purification and characterization of an antimicrobial peptide, termed plantaricyclin A (PlcA), produced by a Lactobacillus plantarum strain isolated from olives. This peptide has a circular structure, and all genes involved in its production, circularization, and secretion were identified. PlcA shows antimicrobial activity against different strains, including Alicyclobacillus acidoterrestris, a common spoilage bacterium, which causes substantial economic losses in the beverage industry every year. In this study, we describe a circular antimicrobial peptide, PlcA, for a Lactobacillus plantarum strain.

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Purification and Partial Characterization of Bacteriocin Lac-B23, a Novel Bacteriocin Production by Lactobacillus plantarum J23, Isolated From Chinese Traditional Fermented Milk.

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