Production of three anti-listerial peptides by Lactobacillus curvatus in MRS broth

Three novel bioactive peptides (BAP1–3) from Lactobacillus curvatus CWBI-B28 were isolated and purified using de Man, Rogosa and Sharp (MRS) broth by a three-step protein purification protocol including ammonium sulfate precipitation, hydrophobic C₁₈ Sep-Pak column and reverse-phase high performance liquid chromatography (RP-HPLC). This procedure allowed the recovery of chromatographically pure antimicrobial peptides with the yields of 19%, 10% and 15% of BAP1, BAP2 and BAP3, respectively. The respective apparent molecular masses as determined by tricine-SDS-polyacrilamide gel electrophoresis were 6365, 3426 and 3496. N-terminal amino acid sequencing of the BAPs and comparison of their sequences with those of international data bases indicated that BAP1 is more likely to be a casein-derived bioactive protein produced upon hydrolysis of the tryptone present in MRS broth by Lb. curvatus CWBI-B28 during active growth. However, the identity of BAP2 and BAP3 could not be determined with certainty; yet, they would be novel bacteriocins not fitting in any of the known classes of bacteriocins. Therefore, this strain would have the ability to produce intrinsic antimicrobial substances and also release bioprotective peptides from milk-proteins upon cultivation in milk or casein-containing food products due to its proteolytic activity. Thus, Lb. curratus CWBI-B28 possesses a good potential to be used in food preservation and safety.