[Multidimensional metabolic engineering of Yarrowia lipolytica for β-carotene production].

Abstract

β-carotene is a natural lipid-soluble carotenoid widely used in the food and pharmaceutical industries. Although metabolic engineering efforts for the production of β-carotene via microbial fermentation have achieved certain progress, there remains room for further improvement. Compared with Saccharomyces cerevisiae or Escherichia coli, Yarrowia lipolytica exhibits a pronounced lipid-flux orientation in its core metabolic logic, conferring a natural advantage for the production of lipid-soluble compounds. This study aims to engineer Y. lipolytica strains for efficient β-carotene production. First, heterologous synthesis of β-carotene was achieved by replacing native promoters with strong ones, attenuating competing pathways, and introducing a bifunctional enzyme derived from Mucor circinelloides. Subsequently, β-carotene production was enhanced through morphological engineering to inhibit hyphal formation, optimization of key gene copy numbers, and improvement of acetyl-CoA supply. The resulting engineered strain LY20 achieved a β-carotene titer of 346.2 mg/L in shake-flask fermentation. Furthermore, in fed-batch fermentation with glucose as the carbon source in a 7.5-L bioreactor, the β-carotene titer reached 3.56 g/L. This study systematically optimizes the biosynthetic pathway for β-carotene through systems metabolic engineering, highlighting the potential of Y. lipolytica for the engineered synthesis of highly hydrophobic natural products and providing guidance for the efficient production of other lipid-soluble terpenoids.