A cryptic tetronate biosynthetic pathway was identified in Kitasatospora niigatensis DSM 44781 via heterologous expression. Distinct from the currently known biosynthetic pathways, this system utilizes a partially functional nonribosomal peptide synthetase and a broadly selective polyketide synthase to direct the assembly and lactonization of the tetronate scaffold. By employing a permissive crotonyl-CoA reductase/carboxylase to provide different extender units, seven new tetronates (kitaniitetronins A-G) were obtained via precursor-directed biosynthesis.