title: Partial purification and characterization of bacteriocin produced by Enterococcus faecalis DU10 and its probiotic attributes.
abstract: A novel bacteriocin produced by avian duck isolated lactic acid bacterium Enterococcus faecalis DU10 was isolated. This bacteriocin showed a broad spectrum of antibacterial activity against important food-borne pathogens and was purified by size exclusion chromatography followed by reverse-phase high-performance liquid chromatography in a C-18 column. Tricine-SDS PAGE revealed the presence of a band with an estimated molecular mass of 6.3 kDa. The zymogram clearly linked the antimicrobial activity with this band. This result was further confirmed by mass-assisted laser desorption ionization time-of-flight mass spectrometry, since a sharp peak corresponding to 6.313 kDa was detected and the functional groups were revealed by Fourier transform infrared spectroscopy. Bacteriocin DU10 activity was found sensitive to proteinase-K and pepsin and partially affected by trypsin and α-chymotrypsin. The activity of bacteriocin DU10 was partially resistant to heat treatments ranging from 30 to 90°C for 30 min. It also withstood a treatment at 121°C for 10 min. Cytotoxicity of bacteriocin DU10 by methyl-thiazolyl-diphenyl-tetrazolium bromide assay showed that the viability of HT-29 and HeLa cells decreased 60 ± 0.7% and 43 ± 4.8%, respectively, in the presence of 3,200 AU/mL of bacteriocin. The strain withstood 0.3% w/v of bile oxgall and pH 2 affected the bacterial growth between 2 and 4 hr of incubation. Adhesion properties examined with HT-29 cell line showed 69.85% initial population of strain E. faecalis DU10, which was found to be strongly adhered to this cell line. These results conclude bacteriocin DU10 may be used as a potential biopreservative and E. faecalis DU10 may be used as a potential probiont to control Salmonella infections.
authors.affiliation: a Department of Biotechnology, School of Life Sciences , Pondicherry University , Puducherry , India.
authors.forename: Venkatesh
authors.initials: V
authors.lastname: Perumal
authors.affiliation: a Department of Biotechnology, School of Life Sciences , Pondicherry University , Puducherry , India.
authors.forename: Ayyanna
authors.initials: A
authors.lastname: Repally
authors.affiliation: a Department of Biotechnology, School of Life Sciences , Pondicherry University , Puducherry , India.
authors.forename: Ankaiah
authors.initials: A
authors.lastname: Dasari
authors.affiliation: a Department of Biotechnology, School of Life Sciences , Pondicherry University , Puducherry , India.
authors.forename: Arul
authors.initials: A
authors.lastname: Venkatesan
mesh_terms: D001422:Bacterial Adhesion; D001430:Bacteriocins / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification*; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D013293:Enterococcus faecalis / Q000378:metabolism*; D019073:HT29 Cells; D006801:Humans; D008970:Molecular Weight; D019936:Probiotics; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D001430:Bacteriocins
keywords: Bacteriocins; E. faecalis DU10; MTT assay; cell adhesion; cytotoxicity; probiotics