Bacteriocin Produced by Lactobacillus crispatus KLB46 Isolated from Human Vagina

초록

Lactobacilli have been considered to play an important role in the health of human vagina. They secrete inhibitory substances to prevent vaginal infection by other organisms. In a previous study, we isolated lactobacilli from Korean women and one of them (KLB46) was identified as Latobacillus crispatus by 16S rDNA sequencing. In this study, we found that L. crispatus KLB46 produced bacteriocin which was highly heat resistant. The maximum production of the bacteriocin was found to occur during the stationary phase of the growth. From plasmid curing experiment we found that plasmid was not involved in the bacteriocin production. In order to characterize the bacteriocin, we carried out a series of purifications. L. crispatus KLB46 was grown for 15 h and removed by centrifugation and the bacteriocin present in the supernatant fraction was concentrated by adding 80% ammonium sulfate. The precipitate pelleted by centrifugation was dissolved in 20mM sodium phosphate buffer, dialyzed against same buffer, and ultra-filtered. The concentrated sample was applied to a CM cation-exchange column before being eluted in 20mM sodium phosphate buffer with 1M NaCl. Fractions showing antimicrobial activity were collected and applied to an SDS-PAGE to estimate molecular mass. Taken together, it is likely that two distinct peptide components having molecular masses ranging 4 Kd and 6.5 Kd should be combined to have the antimicrobial activity.