A Novel Two-Component Bacteriocin, Acidicin P, and Its Key Residues for Inhibiting Listeria monocytogenes by Targeting the Cell Membrane

Listeria monocytogenes is an important pathogen which easily contaminates food and causes fatal systemic infections in human. Bacteriocins have received much attention regarding their natural methods of controlling health-related pathogens. Here, we investigated and characterized a novel two-component bacteriocin named acidicin P from Pediococcus acidilactici LAC5-17. Acidicin P showed obvious antimicrobial activity to L. monocytogenes. Through a sequence similarity network analysis for two-component bacteriocin precursors mined in the RefSeq database, acidicin P was observed to belong to an unusual group of two-component bacteriocins. Acidicin P contains two peptides designated Adpα and Adpβ which are assessed to interact with each other and form a helical dimer structure which can be inserted into the lipid bilayer of target cell membrane. We demonstrate that A5, N7, and G9 in the A(5)xxxG(9) motif of Adpα and S16, R19, and G20 in the S(16)xxxG(20) motif of Adpβ played crucial roles in stabilizing the helix-helix interaction of Adpα and Adpβ and were essential for the antilisterial activity of acidicin P by site-directed mutagenesis. A positive residue, R14, in Adpα and a negative residue, D12, in Adpβ are also important for acidicin P to fight against L. monocytogenes. These key residues are supposed to form hydrogen bonding, which is crucial for the interaction of Adpα and Adpβ. Furthermore, acidicin P induces severe permeabilization and depolarization of the cytoplasmic membrane and causes dramatic changes in L. monocytogenes cell morphology and ultrastructure. Acidicin P has the potential to be applied to inhibit L. monocytogenes efficiently both in the food industry and medical treatments. IMPORTANCE L. monocytogenes can cause widespread food contamination and severe human listeriosis, which amount to a large proportion of the public health and economic burdens. Today, L. monocytogenes is usually treated with chemical compounds in the food industry or antibiotics for human listeriosis. Natural and safe antilisterial agents are urgently required. Bacteriocins are natural antimicrobial peptides that have comparable narrow antimicrobial spectra and are attractive potentials for precision therapy for pathogen infection. In this work, we discover a novel two-component bacteriocin designated acidicin P, which shows obvious antilisterial activity. We also identify the key residues in both peptides of acidicin P and demonstrate that acidicin P is inserted into the target cell membrane and disrupts the cell envelop to inhibit the growth of L. monocytogenes. We believe that acidicin P is a promising lead for further development as an antilisterial drug.