De Novo High-Titer Biosynthesis of (−)-α-Bisabolol in Engineered Candida tropicalis via Green Metabolic Engineered Strategies

(−)-α-bisabolol, a valuable sesquiterpene with anti-inflammatory and antibacterial properties, was sustainably produced via de novo high-titer biosynthesis in Candida tropicalis using engineered cell factories. To minimize resource-intensive extraction methods, we employed green metabolic engineering strategies, including an optimized Scaffold-Mediated Assembly Regulation Tool for coexpressing MrBBS and ERG20 genes, enhancing (−)-α-bisabolol titer 3.24-fold over the control strain. A novel Linker- and Scaffold-Mediated Assembly Regulation Tool further improved pathway efficiency by coordinating eight key acetyl-CoA/mevalonate pathway genes, achieving a 5.23-fold increase. Sustainable strain optimization included knockout of the competing gene DPP1 (yielding 261.2 mg•L–1) and the peroxisomal surface display of Cat2 and ERG10 (290.1 mg•L–1). Additional deletions of genes ADH1 and GPD1 reduced metabolic waste, boosting the titer 2.46-fold. A low-energy two-stage temperature control strategy further improved the titer to 1108.2 mg•L–1, while fed-batch fermentation in a 5 L fermenter─optimized for minimal energy and substrate input─achieved a record titer of 36.9 g•L–1. This study demonstrated a green, scalable biosynthesis platform for (−)-α-bisabolol, reducing reliance on petrochemical or plant-extraction routes, and provided a sustainable blueprint for terpenoid production in nonconventional yeasts.