An Exported Inducer Peptide Regulates Bacteriocin Production in Enterococcus faecium CTC492

Abstract

Production of the bacteriocins enterocin A and enterocin B inEnterococcus faecium CTC492 was dependent on the presence of an extracellular peptide produced by the strain itself. This induction factor (EntF) was purified, and amino acid sequencing combined with DNA sequencing of the corresponding gene identified it as a peptide of 25 amino acids. The gene encodes a prepeptide of 41 amino acids, including a 16-amino-acid leader peptide of the double-glycine type. Environmental factors influenced the level of bacteriocin production in E. faecium CTC492. The optimal pH for bacteriocin production was 6.2. At pH 5.5, growth was slow, and very little bacteriocin was formed. The presence of NaCl or ethanol (EtOH) was also inhibitory to bacteriocin production, and at high concentrations of these solutes, no bacteriocin production was observed. The induction factor induced its own synthesis, and by dilution of the culture 106 times or more, nonproducing cultures were obtained. Bacteriocin production was induced in these cultures by addition of EntF. The response was linear, and low bacteriocin production could be induced by about 10−17 M EntF. This response was attenuated by low pH or the presence of high concentrations of NaCl or EtOH, and 300 times more EntF was needed to induce detectable bacteriocin production in the presence of 6.5% NaCl. High levels of bacteriocin production in cultures grown at low pH or in the presence of high concentrations of NaCl or EtOH were obtained by addition of sufficient amounts of EntF.

Diversity of enterococcal bacteriocins and their grouping in a new classification scheme

“…It encodes a prepeptide with a characteristic double-glycine-type leader peptide of 16 amino acids and mature EntF consists of 25 amino acids. However, O' Keeffe et al (1999) reported a longer, 23-amino-acid leader peptide for EntF, and suggested that the 16-amino-acid leader peptide reported by Nilsen et al (1998) was erroneous and may be the result of a sequencing error. A chemically synthesized, mature EntF peptide induced bacteriocin production by E. faecium CTC492 and DPC1146 that no longer produced bacteriocin as a result of dilution in liquid medium (Nilsen et al, 1998;O'Keeffe et al, 1999), confirming its role as an induction factor.…”

“…However, O' Keeffe et al (1999) reported a longer, 23-amino-acid leader peptide for EntF, and suggested that the 16-amino-acid leader peptide reported by Nilsen et al (1998) was erroneous and may be the result of a sequencing error. A chemically synthesized, mature EntF peptide induced bacteriocin production by E. faecium CTC492 and DPC1146 that no longer produced bacteriocin as a result of dilution in liquid medium (Nilsen et al, 1998;O'Keeffe et al, 1999), confirming its role as an induction factor. The double-glycine-type leader peptide for EntA indicates that ABC transporter gene (entT) and accessory gene (entD) products comprise a dedicated transport system for this bacteriocin (O'Keeffe et al, 1999).…”

Class IIa bacteriocins: biosynthesis, structure and activity

“…4) [52,53]. Like the corresponding class IIa bacteriocins, the IFs are small, heat-stable, cationic and hydrophobic peptides that are ¢rst synthesized as prepeptides with leader sequences of the double-glycine type [4,54]. A three-component system (IF, HPK and RR) has been shown to be responsible for production of most regulated class IIa bacteriocins [4,43,48,54^56].…”

“…It is suggested that the three-component system could be ¢rst triggered as a consequence of an excess in IF concentration occurring through slow accumulation, as a consequence of cell growth [4]. Within such a mechanism, the IF may function as a cell density signal, which seems to be largely reported [4,44,54,55]. Some authors however sustain the idea that signal-transducing system for class IIa bacteriocins, may mediate the response to an environmental signal [43,48].…”

“…In fact, LAB are known to be more or less capable of producing bacteriocins depending on environmental conditions [58^61], which could possibly be connected to the bacteriocin regulatory system. Nilsen et al [54] have suggested that environmental factors may a¡ect the binding of the IF to the HPK or the balance of phosphorylation required for the activation of the RR. Nevertheless, still little or nothing is known about the mechanisms and the extent of implication of environmental factors in this system.…”