De novo biosynthesis of cannabinoid and its analogs in Yarrowia lipolytica

Cannabis sativa has long been a cornerstone of both medicinal and cultural practices, with its therapeutic use spanning over 2,700 years. Central to its therapeutic effects are cannabinoids, which interact with the endocannabinoid system to influence various physiological processes such as anxiety, pain, and inflammation. Despite its benefits, cannabinoid production faces challenges and scarcity from plant extraction. This work leverages Yarrowia lipolytica as a platform for cannabinoid biosynthesis. By optimizing precursor supply, engineering biomolecular condensate-like dual prenyltransferase expression and expanding endogenous metabolism with a noncanonical polyketide synthase, we achieved the de novo biosynthesis of various cannabinoid and its analogs. Our engineered Y. lipolytica produced ~3.5 mg/L of cannabigerolic acid, 18.8 mg/L of orsellinic acid, and 0.5 mg/L of cannabigerorcinic acid. Additionally, CBGA titer reached 15.7 mg/L with olivetolic acid supplement. This work demonstrates Y. lipolytica's versatility as a promising host for cannabinoid and its analogs production, which opens avenues for further research and medicinal applications.