Engineering amino acid-derived malonyl-CoA pathways to boost polyketide production in Yarrowia lipolytica

Abstract Malonyl-CoA is a central precursor involved in the synthesis of various bio-based chemicals, including polyketides, fatty acids, and flavonoids. However, the production of these chemicals is often limited by the availability of malonyl-CoA. Based on retrosynthesis principles, we designed two thermodynamically favorable malonyl-CoA pathways using L-glutamate and L-aspartate as substrates. The novel pathways leverage oxidative deamination and decarboxylation reactions and efficiently channel metabolic flux toward malonyl-CoA, resulting in increased production of total polyketides beyond the capacity of the native acetyl-CoA carboxylase route using glucose as substrate. We also discovered a new-to-nature polyketide (4-hydroxy-6-hydroxyethyl-2-pyrone) derived from the side activity of the TAL pathway, reaching 6.4 g/L in Y. lipolytica . This work highlights the utility of the novel malonyl-CoA pathways in enhancing polyketide production, and the possibility of upcycling abundant amino acids or protein waste in the animal farming or meat industry to produce high-value nonnatural polyketides. Highlights Two thermodynamically favorable malonyl-CoA pathways in Yarrowia lipolytica using L-glutamate and L-aspartate as substrates were engineered. The novel oxygen-dependent malonyl-CoA pathways outperformed the canonical acetyl-CoA carboxylase pathway in polyketide production. A new-to-nature polyketide, 4-hydroxy-6-hydroxyethyl-2-pyrone (HHEP), was produced at a remarkably high titer of 6.4 g/L in shaking flask. The engineered pathway improved the total production of malonyl-CoA-derived compounds (TAL, HHEP, and lipids).