Abstract:[Objective] A genome-scale metabolic model (GSMM) of the high L-malic acid-producing strain Aspergillus oryzae (A. oryzae) FMME-S-38 was constructed and analyzed. [Method] The new GSMM iJW1333 was established for A. oryzae FMME-S-38 through a two-step process of draft reconstruction and model refinement. [Result] The model integrated 1 723 biochemical reactions, 1 723 metabolites, and 1 333 functional genes, achieving the prediction accuracy of 94.6% for substrate metabolism and 92.0% for cell growth. Comparative genomic analysis revealed that mutations in the genes involved in the hexose monophosphate pathway (HMP) and oxidative phosphorylation underpinned the enhanced L-malic acid production of A. oryzae FMME-S-38. Nitrogen source and cofactors (NADH and ATP) were identified as key determinants of efficient L-malic acid biosynthesis. Reducing the mass concentration of nitrogen source and the level of dissolved oxygen (DO) attenuated HMP while strengthening the reductive tricarboxylic acid (rTCA) branch, enabling A. oryzae FMME-S-38 to achieve an L-malic acid titer of (154.2±1.8) g/L.[Conclusion] This study clarifies the critical pathways and physiological mechanisms responsible for high L-malic acid production, providing a solid foundation for employing A. oryzae FMME-S-38 in the biosynthesis of other valuable metabolites.
