Structural Basis of Sequential Enantioselective Epoxidation by a Flavin‐Dependent Monooxygenase in Lasalocid A Biosynthesis

Polyether polyketides are a structurally diverse group of natural products known for their antimicrobial and antiproliferative activities. Lasalocid A is a canonical natural polyether produced by the soil bacterium Streptomyces lasalocidi. In lasalocid A biosynthesis, a polyene polyketide intermediate is converted into a bisepoxide by the flavin-dependent monooxygenase enzyme Lsd18. Remarkably, Lsd18 acts on two distinct C═C groups in the substrate molecule, forming two (R,R) epoxides. We have determined the X-ray crystal structures of Lsd18 in the substrate-free, substrate-bound, and product-bound forms. Our work has revealed that Lsd18 has an extra-large substrate-binding pocket that allows the polyene to adopt different conformations within the enzyme pocket. This feature enables Lsd18 to epoxidate both of the C═C groups. Additionally, a subpocket located near the Lsd18 active site controls stereoselectivity by dictating which face of the C═C group is placed next to the flavin. Molecular understanding of how Lsd18 transforms a polyene into a bisepoxide during lasalocid A biosynthesis lays the foundation for the production of designer polyethers for drug development.