Improving nisin production by increasing nisin immunity/resistance genes in the producer organism Lactococcus lactis

Kim, W. S.; Hall, Robert J.; Dunn, Noel W.

doi:10.1007/s002530051316

Cited by 75 publications

(36 citation statements)

References 24 publications

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“…Even with a high number of copies (8), the level of production by pGW131 and pGW133 was not higher than that by the wild type. This might be caused by limited regulation and posttranslational modification systems or self-toxicity, which is also a limiting factor for the natural producer to express higher levels of bacteriocin, despite the fact that the immunity protein usually protects the producer up to a certain level (29,30).…”

Section: Discussionmentioning

confidence: 99%

Development of a Homologous Expression System for and Systematic Site-Directed Mutagenesis Analysis of Thurincin H, a Bacteriocin Produced by Bacillus thuringiensis SF361

Wang

Manns

Churey

et al. 2014

Appl Environ Microbiol

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Thurincin H is an antimicrobial peptide produced by Bacillus thuringiensis SF361. With a helical back bone, the 31 amino acids of thurincin H form a hairpin structure maintained by four pairs of very unique sulfur-to-␣-carbon thioether bonds. The production of thurincin H depends on a putative gene cluster containing 10 open reading frames. The gene cluster includes three tandem structural genes (thnA1, thnA2, and thnA3) encoding three identical 40-amino-acid thurincin H prepeptides and seven other genes putatively responsible for prepeptide processing, regulation, modification, exportation, and self-immunity. A homologous thurincin H expression system was developed by transforming a thurincin H-deficient host with a novel expression vector, pGW133. The host, designated B. thuringiensis SF361 ⌬thnA1 ⌬thnA2 ⌬thnA3, was constructed by deletion of the three tandem structural genes from the chromosome of the natural thurincin H producer. The thurincin H expression vector pGW133 was constructed by cloning the thurincin H native promoter, thnA1, and a Cry protein terminator into the Escherichia coli-B. thuringiensis shuttle vector pHT315. Thirty-three different pGW133 variants, each containing a different point mutation in the thnA1 gene, were generated and separately transformed into B. thuringiensis SF361 ⌬thnA1 ⌬thnA2 ⌬thnA3. Those site-directed mutants contained either a single radical or conservative amino acid substitution on the thioether linkage-forming positions or a radical substitution on all other nonalanine amino acids. The bacteriocin activities of B. thuringiensis SF361 ⌬thnA1 ⌬thnA2 ⌬thnA3 carrying different pGW133 variants against three different indicator strains were subsequently compared. Bacteriocins are ribosomally synthesized, antimicrobial peptides or proteins produced by bacteria that usually have a narrow inhibitory spectrum, although notable exceptions exist (1). Bacteriocins produced by lactic acid bacteria (LAB) have been extensively studied and used as natural food preservatives. Due to the fact that LAB have a generally recognized as safe (GRAS) status, they have been investigated as potential agents for preventing spoilage and enhancing the safety of foods. The LAB bacteriocins can be divided into three main classes: the class I lantibiotics, containing a lanthionine thioether bond linking the sulfur atom of cysteines with the ␤ carbon of other amino acids; the class II nonlanthionine-containing bacteriocins; and the class III heat-labile large proteins (2). In recent years, bacteriocins produced by Bacillus spp. have gained increasing research interest since many of them exhibit a broader antimicrobial spectrum than most lactic acid bacteriocins, and potential applications in the food, agricultural, and pharmaceutical industries in controlling various spoilage and pathogenic microorganisms are anticipated (3, 4). One Bacillus bacteriocin, thurincin H, is an antimicrobial peptide produced by Bacillus thuringiensis SF361, a strain originally isolated from U.S. domestic sunflower honey. I...

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Section: Discussionmentioning

confidence: 99%

Development of a Homologous Expression System for and Systematic Site-Directed Mutagenesis Analysis of Thurincin H, a Bacteriocin Produced by Bacillus thuringiensis SF361

Wang

Manns

Churey

et al. 2014

Appl Environ Microbiol

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“…For some bacteriocins, it has been shown that engineering of bacteriocin immunity leads to higher bacteriocin production yields (25,27), showing that self-toxicity poses a burden to increase bacteriocin productivity. Moreover, downregulation of key enzymes involved in sugar metabolism has been shown to be involved in tolerance to antibiotics (4).…”

Section: Discussionmentioning

confidence: 99%

The Lcn972 Bacteriocin-Encoding Plasmid pBL1 Impairs Cellobiose Metabolism in Lactococcus lactis

Campelo

Gaspar

Roces

et al. 2011

Appl Environ Microbiol

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pBL1 is a Lactococcus lactis theta-replicating 10.9-kbp plasmid that encodes the synthetic machinery of the bacteriocin Lcn972. In this work, the transcriptomes of exponentially growing L. lactis strains with and without pBL1 were compared. A discrete response was observed, with a total of 10 genes showing significantly changed expression. Upregulation of the lactococcal oligopeptide uptake (opp) system was observed, which was likely linked to a higher nitrogen demand required for Lcn972 biosynthesis. Strikingly, celB, coding for the membrane porter IIC of the cellobiose phosphoenolpyruvate-dependent phosphotransferase system (PTS), and the upstream gene llmg0186 were downregulated. Growth profiles for L. lactis strains MG1363, MG1363/pBL1, and MG1363 ⌬celB grown in chemically defined medium (CDM) containing cellobiose confirmed slower growth of MG1363/pBL1 and MG1363 ⌬celB, while no differences were observed with growth on glucose. The presence of pBL1 shifted the fermentation products toward a mixed acid profile and promoted substantial changes in intracellular pool sizes for glycolytic intermediates in cells growing on cellobiose as determined by highpressure liquid chromatography (HPLC) and nuclear magnetic resonance (NMR). Overall, these data support the genetic evidence of a constriction in cellobiose uptake. Notably, several cell wall precursors accumulated, while other UDP-activated sugar pools were lower, which could reflect rerouting of precursors toward the production of structural or storage polysaccharides. Moreover, cells growing slowly on cellobiose and those lacking celB were more tolerant to Lcn972 than cellobiose-adapted cells. Thus, downregulation of celB could help to build up a response against the antimicrobial activity of Lcn972, enhancing self-immunity of the producer cells.

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“…Since techniques now available for the sitedirected mutagenesis of bacteriocin structural genes (Kim et al, 1998) and with the help of genomics and proteomics, the possibility of constructing new families of designed peptides with enhanced antimicrobial activity or improved stability and specificity characteristics has become a real possibility.…”

Section: Geneticsmentioning

confidence: 99%

Bacteriocins from lactic acid bacteria: purification, properties and use as biopreservatives

Parada

Caron

Medeiros

et al. 2007

Braz. arch. biol. technol.

281

177

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Improving nisin production by increasing nisin immunity/resistance genes in the producer organism Lactococcus lactis

Cited by 75 publications

References 24 publications

Development of a Homologous Expression System for and Systematic Site-Directed Mutagenesis Analysis of Thurincin H, a Bacteriocin Produced by Bacillus thuringiensis SF361

Development of a Homologous Expression System for and Systematic Site-Directed Mutagenesis Analysis of Thurincin H, a Bacteriocin Produced by Bacillus thuringiensis SF361

The Lcn972 Bacteriocin-Encoding Plasmid pBL1 Impairs Cellobiose Metabolism in Lactococcus lactis

Bacteriocins from lactic acid bacteria: purification, properties and use as biopreservatives

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