Mode of action of lactocin 160, a bacteriocin from vaginal Lactobacillus rhamnosus.
OBJECTIVE
To determine the mechanism of antimicrobial action of lactocin 160, a bacteriocin produced by the healthy vaginal strain of Lactobacillus rhamnosus, using an established model, with Micrococcus luteus ATCC 10420 as a test organism.


METHODS
Sensitivity of M. luteus to lactocin 160 was determined by the diffusion assay. Loss of cellular ATP in the lactocin-treated cells was elucidated using a commercially available ATP determination kit (luciferin-luciferase bioluminescence assay). Luminescence intensity as a reflection of ATP quantity was determined using a luminometer. Dissipation of membrane potential (Deltapsi) was studied using fluorophore DiSC3(5) with the fluorescence spectrum sensitive to changes in Deltapsi.


RESULTS
Lactocin 160 inhibited growth of M. luteus ATCC 10420 at a concentration of 5 microg/ml. There were no significant changes in the intracellular ATP level of M. luteus upon the addition of 20 microg/ml of lactocin 160. However, the extracellular ATP level increased significantly. This means that the treatment of cells with lactocin 160 resulted in an efflux of ATP from inside the cells. Therefore, a partially purified lactocin 160 preparation (16 microg /ml of the bacteriocin in the sample) killed sensitive cells and dissipated 3.12 +/- 0.36% of Deltapsi.


CONCLUSIONS
Lactocin 160 has a mode of action typical for bacteriocins. It disturbs the cellular membrane (Deltapsi dissipation) and induces ATP efflux, most likely because of the pore formation, which is a common mechanism of action for many bacteriocins.
Li
Jie
J
Rutgers, The State University of New Jersey, New Brunswick, NJ 08901-8520, USA.
Aroutcheva
Alla A
AA
Faro
Sebastian
S
Chikindas
Michael L
ML
D000255:Adenosine Triphosphate / Q000032:analysis / Q000378:metabolism; D001430:Bacteriocins / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D006801:Humans; D007778:Lactobacillus / Q000378:metabolism*; D008564:Membrane Potentials / Q000187:drug effects; D008826:Microbial Sensitivity Tests; D016982:Micrococcus luteus / Q000187:drug effects / Q000254:growth & development / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D014621:Vagina / Q000382:microbiology*
D001430:Bacteriocins; D000255:Adenosine Triphosphate