Enhanced Expression of CsCCD2 Enables Highly Efficient Crocetin Production in Yarrowia lipolytica

Crocetin, a bioactive carotenoid derivative from saffron, exhibits antioxidant, anti-inflammatory, and metabolic regulatory properties. Its low natural abundance and limited extraction production hinder large-scale production. Microbial synthesis offers a promising alternative but is constrained by low carotenoid cleavage dioxygenase (CCD2) expression. Here, we engineered a zeaxanthin-producing Yarrowia lipolytica strain to enhance CsCCD2 expression via fermentation temperature optimization, SUMOstar fusion, subcellular localization, and precursor supply improvement. The engineered strain achieved a crocetin titer of 196.52 mg/L in shake flasks and 1735.73 mg/L in 10 L fed-batch fermentation, setting a new microbial production record─16-fold higher than previous reports. This work provides a foundation for efficient, large-scale microbial crocetin production and its industrial application.