Engineering Yarrowia Lipolytica for De Novo Biosynthesis of Abscisic Acid

Abscisic acid (ABA) is a sesquiterpenoid plant hormone widely used in agriculture and the food industry. In Yarrowia lipolytica, ABA production relies on complex heterologous enzymatic pathways that have not been systematically engineered, which severely limits biosynthetic efficiency. Here, we establish de novo ABA biosynthesis in Y. lipolytica. We optimized carbon flux by up- and down-regulating key genes, and rationally engineered rate-limiting cytochrome P450 monooxygenases BcABA2 to improve pathway efficiency. Electron transfer was engineered by incorporating an NADPH regeneration module to enhance BcABA2 catalysis. Peroxisomal compartmentalization of the entire ABA pathway, followed by complementation of the defective tag to obtain a complete strain, increased ABA titer to 330.69 mg/L. The engineered strain was scaled up in a 5 L bioreactor, achieving an ABA titer of 2554.36 mg/L after 168 h, which is the highest titer reported to date for microbial fermentation. This work provides feasible enzyme and metabolic engineering strategies for microbial cell factory production of ABA and other terpenoids.