Characterization of the nisin gene cluster nisABTCIPR of Lactococcus lactis. Requirement of expression of the nisA and nisI genes for development of immunity
TLDR
Transcription analyses of several L. lactis strains indicated that an expression product of the nisA gene, together with NisR, is required for the activation of nisinA transcription, indicating that NisI plays a role in the immunity mechanism.Abstract:
The nisin gene cluster nisABTCIPR of Lactococcus lactis, located on a 10-kbp DNA fragment of the nisin-sucrose transposon Tn5276, was characterized. This fragment was previously shown to direct nisin-A biosynthesis and to contain the nisP and nisR genes, encoding a nisin leader peptidase and a positive regulator, respectively [van der Meer, J. R., Polman, J., Beerthuyzen, M. M., Siezen, R. J., Kuipers, O. P. & de Vos, W. M. (1993) J. Bacteriol. 175, 2578–2588]. Further sequence analysis revealed the presence of four open-reading frames, nisB, nisT, nisC and nisI, downstream of the structural gene nisA. The nisT, nisC and nisI genes were subcloned and expressed individually in Escherichia coli, using the T7-RNA-polymerase system. This resulted in the production of radio-labelled proteins with sizes of 45 kDa (NisC) and 32 kDa (NisI). The nisT gene product was not detected, possibly because of protein instability. The deduced amino acid sequence of NisI contained a consensus Iipoprotein signal sequence, suggesting that this protein is a lipid-modified extracellular membrane-anchored protein. Expression of nisI in L. Iactis provided the cells with a significant level of protection against exogeneously added nisin, indicating that NisI plays a role in the immunity mechanism. In EDTA-treated E. coli cells, expression of nisI conferred up to a 170-fold increase in immunity against nisin A compared to controls. Moreover, a lactococcal strain deficient in nisin-A production, designated NZ9800, was created by gene replacement of nisA by a truncated nisA gene and was 10-fold less resistant to nisin A than the wild-type strain. A wild-type immunity level to nisin and production of nisin was obtained in strain NZ9800 harboring complementing nisA and nisZ plasmids. Transcription analyses of several L. IIactis strains indicated that an expression product of the nisA gene, together with NisR, is required for the activation of nisA transcription.read more
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Specificity and Application of the Lantibiotic Protease NisP.
TL;DR: The results indicate that NisP is a suitable protease for the activation of diverse heterologously expressed lantibiotics, which is required to release active antimicrobial compounds.
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Expression of nisin genes in cheese—A quantitative real-time polymerase chain reaction approach
TL;DR: This study is the first in which the evolution of bacteriocin gene transcripts has been quantified rigorously in a cheese-like medium and correlations were established that contribute to the explanation of regulation of nisin biosynthesis and immunity.
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Molecular Analysis of Expression of the Lantibiotic Pep5 Immunity Phenotype
TL;DR: Pep5 production and the immunity phenotype have been found to be tightly coupled, and homologous and heterologous expression of pepI from a xylose-inducible promoter resulted in significant Pep5 insensitivity.
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Production of Functionalized Biopolyester Granules by Recombinant Lactococcus lactis
TL;DR: Overall, it was shown that recombinant L. lactis can be used to manufacture endotoxin-free poly(3-hydroxybutyrate) beads with surface functionalities that are suitable for biomedical applications.
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The genetics of lantibiotic biosynthesis
TL;DR: A number of genes involved in the biosynthesis of these highly modified peptides have been identified, including genes encoding the precursor peptide, enzymes responsible for specific amino acid modifications, proteases able to remove the leader peptide and regulatory proteins controlling lantibiotic biosynthesis.
References
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