ScienceAsia 30 (2004): 115-122 |doi: 10.2306/scienceasia1513-1874.2004.30.115

The Two-synergistic Peptide Bacteriocin Produced by Enterococcus faecium NKR-5-3 Isolated from Thai Fermented Fish (Pla-ra)

Pongtep Wilaipun,^a* Takeshi Zendo,^b Mathana Sangjindavong,^c Sunee Nitisinprasert,^a Vichien Leelawatcharamas,^a Jiro Nakayama^b and Kenji Sonomoto^b

ABSTRACT: Bacteriocin produced by Enterococcus faecium NKR-5-3 isolated from Thai fermented fish was characterized and purified. According to its physical and chemical properties, E. faecium NKR-5-3 produced heat tolerant bacteriocin with broad spectrum activity against indicator strains. Treatment of bacteriocin in cell-free neutralized supernatant (CFNS) with some proteases eliminated or reduced its activity, suggesting it is proteinaceous. Furthermore, it was stable in 0 to 24% NaCl, heat tolerant at pH 2 to 10 and bactericidal toward Enterococcus faecalis ATCC 19433T. Two antibacterial peptides, named enterocin NKR-5-3A and enterocin NKR-5-3B, were purified from the culture supernatant by four steps of column chromatography. Enterocin NKR-5-3A had the same mass, MW=5,241, and N-terminal amino acid sequence as brochocin A produced by Brochothrix campestris ATCC 43754, suggesting it may be identical to brochocin A. Enterocin NKR-5-3B exhibited MW=6,320, but its N-terminal amino acid sequence could not be determined. When these two purified peptides were mixed in a 1:1 ratio, the bacteriocin activity reached a maximum, which corresponded to 64 times the total calculated bacteriocin activity in the mixture, suggesting their synergism.

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^a Department of Biotechnology, Faculty of Agro-Industry, Kasetsart University, Bangkok 10900, Thailand.
^b Department of Bioscience and Biotechnology, Faculty of Agriculture, Kyushu University, Fukuoka 812-
8581, Japan.
^c Department of Fishery Products, Faculty of Fisheries, Kasetsart University, Bangkok 10900, Thailand.

* Corresponding author, E-mail: ffisptw@ku.ac.th

Received 3 Oct 2003, Accepted 1 May 2004