A bio-based economy has the potential to provide sustainable substitutes for petroleum-based products and new chemical building blocks for advanced materials. We previously engineered Saccharomyces cerevisiae for industrial production of the isoprenoid artemisinic acid for use in antimalarial treatments. Adapting these strains for biosynthesis of other isoprenoids such as β-farnesene (CH), a plant sesquiterpene with versatile industrial applications, is straightforward. However, S. cerevisiae uses a chemically inefficient pathway for isoprenoid biosynthesis, resulting in yield and productivity limitations incompatible with commodity-scale production. Here we use four non-native metabolic reactions to rewire central carbon metabolism in S. cerevisiae, enabling biosynthesis of cytosolic acetyl coenzyme A (acetyl-CoA, the two-carbon isoprenoid precursor) with a reduced ATP requirement, reduced loss of carbon to CO-emitting reactions, and improved pathway redox balance. We show that strains with rewired central metabolism can devote an identical quantity of sugar to farnesene production as control strains, yet produce 25% more farnesene with that sugar while requiring 75% less oxygen. These changes lower feedstock costs and dramatically increase productivity in industrial fermentations which are by necessity oxygen-constrained. Despite altering key regulatory nodes, engineered strains grow robustly under taxing industrial conditions, maintaining stable yield for two weeks in broth that reaches >15% farnesene by volume. This illustrates that rewiring yeast central metabolism is a viable strategy for cost-effective, large-scale production of acetyl-CoA-derived molecules.

a b s t r a c tCyclization steps in biosynthesis of aromatic polyketides are typically directed by specific enzymes known as cyclases. The fourth ring cyclization step that forms the fully aromatic pretetramid during tetracycline biosynthesis has not been resolved. Herein we report in vitro and in vivo studies on the fourth ring cyclization step in biosynthesis of SF2575 and related tetracyclic natural products. We demonstrate that an enzyme belonging to the ATP-dependent acyl-CoA ligase family catalyzes the C1eC18 Claisen condensation step that forms the A ring and yields the linear, tetracyclic aromatic compound that is the precursor to tetracyclic natural products. The enzyme, SsfL2, is well-conserved in all tetracycline biosynthetic gene clusters discovered to date. It is proposed that SsfL2 directly adenylates the tricyclic carboxylic acid to facilitate the final carbon-carbon bond formation, and coenzyme A is not required.