Dissection of the bridging pattern of bovicin HJ50, a lantibiotic containing a characteristic disulfide bridge.

Bovicin HJ50, a lantibiotic produced by Streptococcus bovis HJ50, is featured by the presence of a disulfide bridge. This study described a simplified in vitro synthetic strategy for producing bovicin HJ50 totally based on Escherichia coli expression system. In this strategy termed as Semi-in vitro biosynthesis (SIVB), prepeptide BovA and modification enzyme BovM were co-expressed to generate posttranslationally modified BovA. Then a specific protease BovT150 was employed to remove the leader peptide in vitro and produce biologically active bovicin HJ50. Via SIVB, a series of ring-broken bovicin mutants C13A, C21A, C29A and T10A/C32A were prepared by introducing site-directed mutations into bovA gene. Further, we analyzed the bridging patterns of these mutants through chemical modification and successfully clarified the bridging pattern of bovicin HJ50. The results showed that two thioether bridges exist between Thr8 and Cys13, and Thr10 and Cys32, respectively, and that the disulfide bond bridging Cys21 and 29 is very relevant for the antimicrobial activity of bovicin HJ50. This is the first study that reports the bridging pattern of bovicin HJ50.