Abstract
Lactic acid bacteria produce a variety of antimicrobial peptides known as bacteriocins. Most bacteriocins are understood to kill sensitive bacteria through receptor‐mediated disruptions. Here, we report on the identification of the Lactobacillus plantarum plantaricin EF (PlnEF) receptor. Spontaneous PlnEF‐resistant mutants of the PlnEF‐indicator strain L. plantarum NCIMB 700965 (LP965) were isolated and confirmed to maintain cellular ATP levels in the presence of PlnEF. Genome comparisons resulted in the identification of a single mutated gene annotated as the membrane‐bound, magnesium/cobalt efflux protein CorC. All isolates contained a valine (V) at position 334 instead of a glycine (G) in a cysteine‐β‐synthase domain at the C‐terminal region of CorC. In silico template‐based modeling of this domain indicated that the mutation resides in a loop between two β‐strands. The relationship between PlnEF, CorC, and metal homeostasis was supported by the finding that PlnEF‐resistance was lost when PlnEF was applied together with high concentrations of Mg2+, Co2+, Zn2+, or Cu2+. Lastly, PlnEF sensitivity was increased upon heterologous expression of LP965 corC but not the G334V CorC mutant in the PlnEF‐resistant strain Lactobacillus casei BL23. These results show that PlnEF kills sensitive bacteria by targeting CorC.
Highlights
Lactic acid bacteria (LAB) produce a diverse array of bacteriocins
We identified CorC, a putative magnesium/cobalt exporter, as the receptor for the L. plantarum bacteriocin plantaricin EF (PlnEF)
These findings are in agreement with previous studies on PlnEF showing that this bac‐ teriocin causes cation efflux (Moll et al, 1999)
Summary
Lactic acid bacteria (LAB) produce a diverse array of bacteriocins. Bacteriocins are ribosomally synthesized peptides with bactericidal activity and are frequently most active against species that are highly related to the producer strains (Chikindas, Weeks, Drider, Chistyakov, & Dicks, 2018). The final optical density (as measured by OD600) and growth rates (ANCOVA, F(3, 44) = 1.75, p = 0.17) of all strains was reduced in the presence of 1,000 nM of the un‐ related bacteriocin plantaricin A (PlnA) (Figure 1) when compared to growth without PlnA, thereby showing that PlnEF resistance was specific and not the result of changes in general, stress‐related re‐ sponses to antibacterial peptides. A dose dependency was found such that strain EF.A initiated growth by 20 hr in the presence of supplemental 250 mM MgSO4, but not 500 mM MgSO4 (Figure 4) These results show that PlnEF acts synergistically with metals to inhibit the growth of LP965 and that the G334V mutation is not sufficient to prevent cell damage at high concentrations of MgSO4, CoSO4, CuSO4, and ZnSO4. These results confirm that wild‐type LP965 corC increases sensitivity to PlnEF and that the single G334V amino acid substitution is sufficient to confer resistance
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