Abstract
BackgroundBacteriocin production in the lactic acid bacterium Lactobacillus plantarum C11 is regulated through a quorum sensing based pathway involving two highly homologous response regulators (59% identity and 76% similarity), PlnC as a transcriptional activator and PlnD as a repressor. Previous in vitro studies have shown that both regulators bind, as homodimers, to the same DNA regulatory repeats to exert their regulatory functions. As the genes for these two proteins are located on the same auto-regulatory operon, hence being co-expressed upon gene activation, it is plausible that their opposite functions must somehow be differentially regulated, either in terms of timing and/or binding kinetics, so that their activities do not impair each other in an uncontrolled manner. To understand the nature behind this potential differentiation, we have studied the binding kinetics of the two regulators on five target promoters (PplnA, PplnM, PplnJ, PplnE and PplnG) from the bacteriocin regulon of L. plantarum C11.ResultsBy using surface plasmon resonance spectroscopy we obtained parameters such as association rates, dissociation rates and dissociation constants, showing that the two regulators indeed differ greatly from each other in terms of cooperative binding and binding strength to the different promoters. For instance, cooperativity is very strong for PlnC binding to the promoter of the regulatory operon (PplnA), but not to the promoter of the transport operon (PplnG), while the opposite is seen for PlnD binding to these two promoters. The estimated affinity constants indicate that PlnC can bind to PplnA to activate transcription of the key regulatory operon plnABCD without much interference from PlnD, and that the repressive function of PlnD might act through a different mechanism than repression of the regulatory operon.ConclusionWe have characterised the DNA binding kinetics of the two regulators PlnC and PlnD from the bacteriocin locus in L. plantarum C11. Our data show that PlnC and PlnD, despite their strong homology to each other, differ greatly from each other in terms of binding affinity and cooperativity to the different promoters of the pln regulon.
Highlights
Bacteriocin production in the lactic acid bacterium Lactobacillus plantarum C11 is regulated through a quorum sensing based pathway involving two highly homologous response regulators (59% identity and 76% similarity), PlnC as a transcriptional activator and PlnD as a repressor
Many lactic acid bacteria produce a group of ribosomally synthesised antibacterial peptides, most frequently referred to as bacteriocins, which kill other bacteria by disrupting the integrity of the membrane of target cells leading to leakage of cellular solutes across the membrane and eventually cell death [1,2,3]
Bacteriocin production is regulated through a quorum sensing pathway mediated by a secreted inducer peptide (IP), a transmembrane sensor histidine protein kinase (HPK) and a cytoplasmic response regulator (RR) [4,5]
Summary
Bacteriocin production in the lactic acid bacterium Lactobacillus plantarum C11 is regulated through a quorum sensing based pathway involving two highly homologous response regulators (59% identity and 76% similarity), PlnC as a transcriptional activator and PlnD as a repressor. Most bacteriocins have quite narrow inhibitory spectra, normally composed of species closely related to the bacteriocin producers. When the critical threshold level of IP is achieved, this signal is processed by the IPbinding protein HPK and subsequently transferred to its cognate RR via a series of phosphorylation reactions that eventually results in a phosphorylated RR. The latter binds as dimers to regulated promoters and triggers expression of all genes involved in bacteriocin biosynthesis [5,9,10,11]
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