Lipidation Engineering in Daptomycin Biosynthesis

Lipopeptides are an important family of natural products, some of which are clinically used as antibiotics to treat multidrug-resistant pathogens. Although the lipid moieties play a crucial role in balancing antibacterial activity and hemolytic toxicity, modifying the lipid moieties has been challenging due to the complexity of the lipidation process in lipopeptide biosynthesis. Here, we show that the lipid profile can be altered by engineering both secondary and primary metabolisms, using daptomycin as an example. First, swapping the fatty acyl AMP ligase (FAAL) gene <i>dptF</i> with foreign FAAL homologs improved the fatty acyl specificity of the lipidation process for decanoic acid. Then, the introduction of <i>Mycobacterium</i> type I fatty acid synthase operon (MvFAS-Ib/MvAcpS) and <i>Cryptosporidium</i> thioesterase (CpTEII) enriched the fatty acid pool with decanoic acid in <i>Streptomyces roseosporus</i>. The engineered fatty acid metabolism eliminates the need for external decanoic acid supplementation by enabling <i>S. roseosporus</i> to biosynthesize decanoic acid. By complete engineering of the lipidation process, we achieved, for the first time, high-purity, natural production of daptomycin. The lipidation engineering approach we demonstrate here lays the foundation for the lipidation control in lipopeptide biosynthesis.