Fermalogic recently discovered that the key erythromycin precursor, methylmalonyl- CoA, is the limiting factor for improved erythromycin production. As such, the methylmalonyl-CoA metabolic node has become a focal point for strain improvement strategies. The main goals of this study are to identify key enzymes and metabolic pathways either known or predicted to affect carbon flow at the methylmalonyl-CoA node and manipulate these genes and pathways for increased erythromycin production. Many other natural products besides erythromycin are made from methylmalonyl-CoA precursors, therefore the information gained from this study could be applicable to a broad range of pharmaceutical compounds including other antibiotics, anti-cancer com- pounds, and immunosuppressants. For the last fifty years strain improvement has been performed by a random mutate-and-screen process that has not generated any information regarding the genetics or biochemistry of strain improvement. This study will pro- vide important information and technology that can be used to rationally engineer Actinomycetes for rapid commercial development of critically needed new natural products, and to improve the production of existing products to reduce the cost of production. Metabolic engineering has as its primary goal the rational application of genetic and biochemical data for the improvement of a production process. The ability to identify important genes, enzymes, and metabolic pathways for an important production process will allow metabolic engineers to impart rational manipulations to the producing strain for improved production. These applications will have far-reaching effects in disciplines such as pharmaceutical production, agriculture, and many other biotechnology applications. The technology that will be developed in this project will elucidate path- ways leading to the biosynthesis of commercially important antibiotics with a focus on erythromycin
