The Inhibitory Spectrum of Thermophilin 9 from Streptococcus thermophilus LMD-9 Depends on the Production of Multiple Peptides and the Activity of BlpG St , a Thiol-Disulfide Oxidase

Abstract

ABSTRACT The blp St cluster of Streptococcus thermophilus LMD-9 was recently shown to contain all the genetic information required for the production of bacteriocins active against other S. thermophilus strains. In this study, we further investigated the antimicrobial activity of S. thermophilus LMD-9 by testing the susceptibility of 31 bacterial species (87 strains). We showed that LMD-9 displays an inhibitory spectrum targeted toward related gram-positive bacteria, including pathogens such as Listeria monocytogenes . Using deletion mutants, we investigated the contribution of the three putative bacteriocin-encoding operons blpD St - orf2 , blpU St - orf3 , and blpE St - blpF St ( bac St operons) and of the blpG St gene, which encodes a putative modification protein, to the inhibitory spectrum and immunity of strain LMD-9. Our results present evidence that the blp St locus encodes a multipeptide bacteriocin system called thermophilin 9. Among the four class II bacteriocin-like peptides encoded within the bac St operons, BlpD St alone was sufficient to inhibit the growth of most thermophilin 9-sensitive species. The blpD St gene forms an operon with its associated immunity gene(s), and this functional bacteriocin/immunity module could easily be transferred to Lactococcus lactis . The remaining three Bac St peptides, BlpU St , BlpE St , and BlpF St , confer poor antimicrobial activity but act as enhancers of the antagonistic activity of thermophilin 9 by an unknown mechanism. The blpG St gene was also shown to be specifically required for the antilisteria activity of thermophilin 9, since its deletion abolished the sensitivities of most Listeria species. By complementation of the motility deficiency of Escherichia coli dsbA , we showed that blpG St encodes a functional thiol-disulfide oxidase, suggesting an important role for disulfide bridges within thermophilin 9.