Discovery, biosynthesis, and characterization of a lanthipeptide from Bacillus subtilis EH11 with a unique lanthionine ring pattern

Lanthipeptide biosynthetic genes are present in a wide range of bacteria, providing convenient tools for engineering bioactive lanthipeptides. Here, we report a class I lanthipeptide biosynthetic gene cluster (lanBTC) in a Bacillus strain, involved in the biosynthesis of a novel lanthipeptide that we termed balucin. Balucin was active against food-borne pathogens such as Bacillus cereus and Listeria monocytogenes. The unusual structural features of balucin showed the presence of aspartic acid residues C-terminally flanking the lanthionine rings. Such negatively charged amino acids directly flanking modifiable cysteines at the C-terminal side have not been reported for precursors that are modified by other LanBCs. Heterologous functional expression of balucin in a dedicated E. coli expression system was achieved, and the protease responsible for cleaving the balucin leader peptide was identified. With this system, the balucin biosynthetic enzymes were successfully employed to modify antimicrobial peptides that have negatively charged residues C-terminally flanking lanthionine-forming cysteines.