Purification of the bacteriocin bavaricin MN and characterization of its mode of action against Listeria monocytogenes scott A cells and lipid vesicles

Abstract

Bavaricin MN was purified from Lactobacillus sake culture supernatant 135-fold with a final yield of 11%. Sequence analysis revealed bavaricin MN to be a 42-amino-acid peptide having a molecular weight of 4,769 and a calculated pI of 10.0. Computer analysis indicated that the C-terminal region may form an α-helical structure with an amphipathic nature deemed important in the interaction of bacteriocins with biological membranes. Bavaricin MN rapidly depleted the membrane potential (Δp) of energized Listeria monocytogenes cells in a concentration-dependent fashion. At a bavaricin MN concentration of 9.0 μg/ml, the Δp decreased by 85%. Both the electrical potential (ΔΨ) and ZΔpH components of the Δp were depleted, and this depletion was not dependent on a threshold level of proton motive force. In addition to studying the effect of bavaricin MN on the Δp of vegetative cells, bavaricin MN-induced carboxyflourescein (CF) efflux from L. monocytogenes-derived lipid vesicles was also characterized. Bavaricin MN- induced CF leakage was also concentration dependent with an optimum of pH 6.0. The rate of CF efflux was 63% greater in lipid vesicles in which a ΔΨ was generated compared with that in lipid vesicles in the absence of ΔΨ.