Insights into the Biosynthesis of Duramycin

Lantibiotics are ribosomally synthesized and posttranslationally modified antimicrobial peptides that are characterized by the thioether cross-linked bisamino acids lanthionine (Lan) and methyllanthionine (MeLan). Duramycin contains 19 amino acids, including one Lan and two MeLans, an unusual lysinoalanine (Lal) bridge formed from the ε-amino group of lysine 19 and a serine residue at position 6, and anerythro -3-hydroxy-l -aspartic acid at position 15. These modifications are important for the interactions of duramycin with its biological target, phosphatidylethanolamine (PE). Based on the binding affinity and specificity for PE, duramycin has been investigated as a potential therapeutic, as a molecular probe to investigate the role and localization of PE in biological systems, and to block viral entry into mammalian cells. In this study, we identified the duramycin biosynthetic gene cluster by genome sequencing ofStreptomyces cinnamoneus ATCC 12686 and investigated thedur biosynthetic machinery by heterologous expression inEscherichia coli . In addition, the analog duramycin C, containing six amino acid changes compared to duramycin, was successfully generated inE. coli . The substrate recognition motif of DurX, an α-ketoglutarate/iron(II)-dependent hydroxylase that carries out the hydroxylation of aspartate 15 of the precursor peptide DurA, was also investigated using mutagenesis of the DurA peptide. Bothin vivo andin vitro results demonstrated that Gly16 is important for DurX activity.IMPORTANCE Duramycin is a natural product produced by certain bacteria that binds to phosphatidylethanolamine (PE). Because PE is involved in many cellular processes, duramycin is an antibiotic that kills bacteria, but it has also been used as a molecular probe to detect PE and monitor its localization in mammalian cells and even whole organisms, and it was recently shown to display broad-spectrum inhibition of viral entry into host cells. In addition, the molecule has been evaluated as treatment for cystic fibrosis. We report here the genes that are involved in duramycin biosynthesis, and we produced duramycin by expressing those genes inEscherichia coli . We show that duramycin analogs can also be produced. The ability to access duramycin and analogs by production inE. coli opens opportunities to improve duramycin as an antibiotic, PE probe, antiviral, or cystic fibrosis therapeutic.