Lipid-donor-anchored genome mining uncovers dioxanopeptins, antibacterial lipopeptides with a 1,3-dioxane functionalized polyunsaturated lipid tail

, and their antibacterial phenotype correlated with disruption of proton motive force homeostasis and ATP production. Removal of the pyruvyl-derived ketal markedly diminished potency, highlighting its contribution to antibacterial activity. Together, these results expand the chemical space of lipopeptides and illustrate how lipid-donor-anchored genome mining can complement NRPS-focused strategies to access functionally differentiated antibiotic scaffolds.