Phoslactomycin (Plm) and fostriecin are members of a group of potent and selective inhibitors of serine-threonine phosphatase 2A (PP2A).  These polyketide natural products all contain an α,β-unsaturated–δ-lactone which forms a covalent adduct with Cys269 which is unique to PP2A.  We have cloned the Plm biosynthetic gene cluster and carried out a detailed analysis of this and the fostriecin biosynthetic gene cluster.  Through a series of gene deletion and complementation studies we have demonstrated that a new post polyketide synthase decarboxylative-dehydration step establishes this critical electrophile.   These antibiotics also contain a linear unsaturated chain containing both cis (Z) and trans (E) double bonds.  A series of studies have provided the first experimental evidence that individual modules in a type I polyketide synthase are responsible for controlling the stereochemistry of the double bond formation.   We have also generated a series of blocked mutants that produce intermediates in the Plm pathway.  Enzyme-catalyzed esterification of one of these intermediates has generated a series of new Plms.  Directed biosynthesis, and combinatorial biosynthesis with the Plm and fostriecin biosynthetic genes have provided an additional array of new Plms with structural diversity across the structure.   The activity of these compounds as PP2A inhibitors will be presented.