Lantibiotics are ribosomally synthesized peptide antibiotics composed of an N-terminal leader peptide that is cleaved to yield the active antibacterial peptide. Significant advancements in molecular tools that promote the study of lantibiotic biosynthesis can be used in Streptococcus mutans. Herein, we further our understanding of leader peptide sequence and core peptide structural requirements for the biosynthesis and transport of the lantibiotic mutacin 1140. Our study on mutacin 1140 biosynthesis shows a dedicated secondary cleavage site within the leader peptide and the dependency of transport on core peptide posttranslational modifications (PTMs). The secondary cleavage site on the leader peptide is found at the ؊9 position, and secondary cleavage occurs before the core peptide is transported out of the cell. The coordinated cleavage at the ؊9 position was absent in a lanT deletion strain, suggesting that the core peptide interaction with the LanT transporter enables uniform cleavage at the ؊9 position. Following transport, the LanP protease was found to be tolerant to a wide variety of amino acid substitutions at the primary leader peptide cleavage site, with the exception of arginine at the ؊1 position. Several leader and core peptide mutations produced core peptide variants that had intermediate stages of PTM enzyme modifications, supporting the concept that PTM enzyme modifications, secondary cleavage, and transport are occurring in a highly coordinated fashion. IMPORTANCEMutacin 1140 belongs to the class I lantibiotic family of ribosomally synthesized and posttranslationally modified peptides (RiPPs). The biosynthesis of mutacin 1140 is a highly efficient process which does not lead to a discernible level of production of partially modified core peptide variants. The products isolated from an extensive mutagenesis study on the leader and core peptides of mutacin 1140 show that the posttranslational modifications (PTMs) on the core peptide occur under a highly coordinated dynamic process. PTMs are dictated by the distance of the core peptide modifiable residues from PTM enzyme active sites. The formation of lanthionine rings aids in the formation of successive PTMs, as was observed in a peptide variant lacking a Cterminal decarboxylation. Lantibiotics are a class of ribosomally synthesized peptide antibiotics produced by Gram-positive bacteria, such as Lactococcus lactis and Streptococcus mutans (1, 2). Lantibiotics are characterized by the presence of posttranslational modifications (PTMs), such as dehydrated residues and lanthionine rings. The modified residues 2,3-didehydroalanine (Dha) and 2,3-didehydrobutyrine (Dhb) are formed from dehydration of serines and threonines, respectively. The cyclization between a cysteine and either a Dha or a Dhb forms a lanthionine or a methyllanthionine ring, respectively. The biosynthetic gene cluster contains all the genes necessary to produce a lantibiotic. The lan operon for class I lantibiotics contains genes encoding the lantibiotic peptide (lanA); the...

Lantibiotics are ribosomally synthesized peptide antibiotics composed of an N-terminal leader peptide that promotes the core peptide's interaction with the post translational modification (PTM) enzymes. Following PTMs, mutacin 1140 is transported out of the cell and the leader peptide is cleaved to yield the antibacterial peptide. Mutacin 1140 leader peptide is structurally unique compared to other class I lantibiotic leader peptides. Herein, we further our understanding of the structural differences of mutacin 1140 leader peptide with regard to other class I leader peptides. We have determined that the length of the leader peptide is important for the biosynthesis of mutacin 1140. We have also determined that mutacin 1140 leader peptide contains a novel four amino acid motif compared to related lantibiotics. PTM enzyme recognition of the leader peptide appears to be evolutionarily distinct from related class I lantibiotics. Our study on mutacin 1140 leader peptide provides a basis for future studies aimed at understanding its interaction with the PTM enzymes.