A Leaderless Two-Peptide Bacteriocin, Enterocin DD14, Is Involved in Its Own Self-Immunity: Evidence and Insights

Enterocin DD14 (EntDD14) is a two-peptide leaderless bacteriocin produced byEnterococcus faecalis14, a strain previously isolated from meconium. EntDD14 has a strong antibacterial activity against Gram-positive bacteria. Leaderless bacteriocins, unlike bacteriocins with leader peptides, are immediately active after their translation, and a producing strain has then to develop specific mechanisms to protect both intra and extracellular compartments. Thein silicoanalysis ofEnt. faecalis14 genome allowed to locate downstream of structuralddABgenes, 8 other adjacent genes, designedddCDEFGHIJ, which collectively may form three operons. To gain insights on immunity mechanisms ofEnt. faecalis14, mutant strains knocked out inddABgenes encoding bacteriocin precursor peptides (Delta bac) and/or ABC transporter (Delta ddI) of EntDD14 were constructed and characterized. Importantly, Delta bacmutant strains, from which structuralddABgenes were deleted, resulted unable to produce EntDD14 and sensitive to exogenous EntDD14 showing their involvement in theEnt. faecalis14 immunity system. Moreover, the sensitivity of Delta bacmutants appeared not to be associated with the down-regulation ofddCDEFGHIJgene expression since they were similarly expressed in both Delta bacand wild-type strains during the log phase while they were found significantly down-regulated in the Delta bacmutant strain after 24 h of growth. Data gathered from this study suggest also the implication of the ABC transporter (ddHIJ) in the active export of EntDD14 but ruled-out its involvement in the primary self-immunity system. Interestingly, non-bacteriocin producingEnt. faecalisJH2-2 cells transformed withddAB, orddABplus genes encoding the ABC transporter (ddAB-HIJ) did not produce EntDD14 and remained sensitive to its action. Of note, trans-complementation of the Delta bacmutant strain with these constructions allowed to recover the WT phenotype. To the best of our knowledge, this is the first study delineating the role of the intracellular two-peptide leaderless bacteriocins in their self-immunity.