Rewiring amino acid metabolism to boost nonnatural polyketide production in Yarrowia lipolytica
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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, leading to increased triacetic acid lactone (TAL) production beyond the capability of the native acetyl-CoA carboxylase. We also discovered a new-to-nature polyketide (4-hydroxy-6-hydroxyethyl-2-pyrone) derived from the side activity of TAL pathway, reaching 5.4 g/L. This work highlights the utility of the novel malonyl-CoA pathways in enhancing polyketide production, and the possibility of using abundant amino acids or protein waste as feedstock to produce high-value nonnatural polyketides.
Significance Statement
We engineered two thermodynamically favorable malonyl-CoA pathways in Yarrowia lipolytica using L-glutamate and L-aspartate as substrates. These pathways circumvent ATP dependency by leveraging oxidative deamination and decarboxylation reactions, offering a thermodynamically-favorable routes for bioproduction from abundant amino acids and waste proteins. The novel pathways outperformed the canonical acetyl-CoA carboxylase pathway in polyketide production, achieving up to moe than 3.7 g/L of triacetic acid lactone and a new-to-nature polyketide, 4-hydroxy-6-hydroxyethyl-2-pyrone (HHEP), at 5.4 g/L in bioreactor. The discovery of HHEP, a structural analog of kojic acid (a strong antimicrobial and skincare ingredient), underscores the potential of microbial cells for generating novel bioactive molecules. Our work highlights the interplay between pathway thermodynamics, enzyme kinetics, and metabolic flux redirection.
