Metabolic model-guided strain design for improved succinic acid production in Yarrowia lipolytica

The oleaginous yeast Yarrowia lipolytica has emerged as a promising microbial host for the production of succinic acid (SA), a key bio-based platform chemical, owing to its metabolic versatility and robustness under industrial fermentation conditions. To enable rational engineering of Y. lipolytica toward enhanced SA production, a genome-scale metabolic model (GEM) of the industrially relevant W29 strain has been reconstructed in this study, comprising 634 genes, 1130 metabolites, and 1364 reactions distributed across eight compartments. The model achieved 88.9 % accuracy in predicting growth phenotypes on 18 carbon sources and demonstrated a robust correlation with experimental growth rates (R² = 0.98). Leveraging in silico strain design tools, a set of knockout and overexpression targets for enhancing SA production from glycerol was identified. Simulations revealed that knocking out succinate dehydrogenase (SDH) and acetyl-CoA hydrolase (ACH) increased SA flux to 4.36 mmol/gDW/h (0.56 g/g glycerol), aligning with prior experimental studies. Overexpression of pyruvate carboxylase and TCA/glyoxylate cycle enzymes was predicted to further enhance SA yields by up to 186 %. These findings not only aligned with experimentally confirmed targets but also uncovered novel interventions, demonstrating the GEM as a robust platform for rational strain design toward enhanced production of SA and other bio-based chemicals.