Bacteriocin T8, a novel class IIa sec-dependent bacteriocin produced by Enterococcus faecium T8, isolated from vaginal secretions of children infected with human immunodeficiency virus

Abstract

Enterococcus faecium T8, isolated from vaginal secretions of children with human immunodeficiency virus, produces a class IIa sec-dependent bacteriocin that is structurally different from three other class IIa sec-dependent bacteriocins, i.e., enterocin P and an enterocin P-like bacteriocin, produced by Enterococcus faecium, and bacteriocin 31, produced by Enterococcus faecalis, and from a class III bacteriocin produced by E. faecalis. The genes encoding the bacteriocin, immunity protein, mobilization protein, and relaxase nuclease are located on a 7-kb plasmid. Bacteriocin T8 has a molecular mass of 5.1 kDa based on its DNA sequence, similar to the 5.0 kDa recorded for bacteriocin 31 but larger than the 4.6 kDa reported for enterocin P. At the amino acid level, bacteriocin T8 is 69% homologous to bacteriocin 31 and 47% homologous to enterocin P. Bacteriocin T8 is active against E. faecalis isolated from patients diagnosed with vaginosis, against Lactobacillus sakei, and against a Propionibacterium sp. The peptide is heat stable (60 min at 100 degrees C) and remains active in phosphate buffer from pH 4.0 to 10.0. The mode of activity is bactericidal, as determined with E. faecalis.

FIG. 1.

Effect of bacteriocin T8 on cell growth of E. faecalis MDK2. Bacteriocin T8 was added to mid-logarithmic-phase cells (after 5 h of growth). Symbols: ▪, growth in the absence of bacteriocin T8; ⧫, growth in the presence of bacteriocin T8. OD, optical density.

FIG. 2.

Nucleotide sequence of the bacteriocin region and deduced amino acid sequence. The −10 promoter box and the Shine-Dalgarno (S.D.) ribosome-binding sequences are underlined. ORF 1 encodes the structural gene of bacteriocin T8, and ORF 2 encodes the potential immunity gene.

FIG. 3.

Hydrophobicity profiles of the leader peptide (A) and the mature bacteriocin (B).

FIG. 4.

Comparison of amino acid sequences of the N termini of class IIa sec-dependent bacteriocins. Data for enterocin P and bacteriocin 31 were obtained from the work of Cintas et al. (5) and Tomita et al. (28), respectively.