Purification and characterization of enterocin 62-6, a two-peptide bacteriocin produced by a vaginal strain ofEnterococcus faecium: Potential significance in bacterial vaginosis

Abstract

A bacteriocin produced by a vaginal isolate of Enterococcus faecium strain 62-6, designated enterocin 62-6, was characterized following purification and DNA sequence analysis and compared to previously described bacteriocins. Enterocin 62-6 was isolated from brain heart infusion (BHI) culture supernatants using ammonium sulfate precipitation followed by elution from a Sepharose cation exchange column using a continuous salt gradient (0.1-0.7 M NaCl). SDS-PAGE of an active column fraction resulted in an electrophoretically pure protein, which corresponded to the growth inhibition of the sensitive Lactobacillus indicator strain in the gel overlay assay. Purified enterocin 62-6 was shown to be heat- and pH-stable, and sensitive to the proteolytic enzymes alpha-chymotrypsin and pepsin. Results from mass spectrometry suggested that it comprised two peptides of 5206 and 5219+/-1 Da, which was confirmed by DNA sequence analysis. The characteristics of enterocin 62-6 as a small, heat- and pH-stable, cationic, hydrophobic, two-peptide, plasmid-borne bacteriocin, with an inhibitory spectrum against a broad range of Gram-positive but not Gram-negative bacteria, were consistent with its classification as a class IIc bacteriocin. Furthermore, its wide spectrum of growth inhibitory activity against Gram-positive bacteria of vaginal origin including lactobacilli, and stability under the acidic conditions of the vagina, are consistent with our hypothesis that it could have potential significance in disrupting the ecology of the vaginal tract and pave the way for the establishment of the abnormal microbiota associated with the vaginal syndrome bacterial vaginosis. This is the first class IIc bacteriocin produced by a strain of E. faecium of vaginal origin to be characterized.

Biochemical characterization and identi cation of enterocins produced by E. hirae And E. faecalis

“…Table 6 shows the effect of various enzymes on inhibitory activity of enterocins .It demonstrated sensitivity to proteolytic enzymes trypsin and  chymotrypsin while treatment with catalase, lipase and lysozyme did not affect the activity of enterocin.Thus,the obtained data suggests that the loss of enterocin activity to proteolytic enzyme is due to the fact that enterocin contained an essential proteinaceous component while stability in presence of both lipase and lysozyme indicates that it lacked lipid or carbohydrate moieties. Similar enterocin activities to different physicochemical parameters were also seen in Cocolin et al,(2007) Our results are also in accordance with the fi ndings of Ahmad et al, (2003) and Dezwaan et al(2007).…”

Enterocin LD3 fromEnterococcus hiraeLD3 causing efflux of intracellular ions and UV‐absorbing materials in Gram‐negative bacteria

“…; Ghrairi and Hani ), whereas enterocin 62‐6 was not able to inhibit the E. coli ATCC 4157 (Dezwaan et al . ). Thus, the inhibition of E. coli cells without pretreatments suggests the unique mode of action of enterocin LD3 and advantage over other bacteriocins.…”

Combined effect of reuterin and lactic acid bacteria bacteriocins on the inactivation of food-borne pathogens in milk

“…However, nisin was the only bacteriocin able to reduce the counts of S. aureus. Nisin and lacticin 481 belong to class I bacteriocins (Dufour, Hindré, Haras, & Pennec, 2007), while enterocin I (Floriano, Ruiz-Barba, & Jiménez-Díaz, 1998) also described as enterocin L50, belongs to class II (Criado et al, 2006;Dezwaan et al, 2007). Class I, characterized by their unusual and posttranslationally modified aminoacids, and class II bacteriocins, are small heat-stable peptides that, because of their abundance and potential application in industrial processes, are the most comprehensively studied.…”