Boosting production of cembratriene‐ol in Saccharomyces cerevisiae via systematic optimization

Abstract Cembratriene‐ol is a good biodegradable biopesticide ingredient with future potential applications in the field of sustainable agriculture. Cembratriene‐ol is a monocyclic diterpenoid compound that is synthesized only in the trichome gland of Nicotiana plants. In this study, geranylgeranyl diphosphate synthase gene ggpps from Taxus canadensis and cbts*Δp were heterologously expressed in Saccharomyces cerevisiae W303‐1A to successfully synthesize cembratriene‐ol. The titer of cembratriene‐ol was increased by 1.84‐fold compared to the control by overexpressing the S. cerevisiae bifunctional (2E,6E)‐farnesyl diphosphate synthase genes ERG20 and cbts*Δp under one promoter P GAP . The titer of cembratriene‐ol in the engineered S. cerevisiae BY4741 was increased by 1.39‐fold compared to the engineered S. cerevisiae W303‐1A. The titer of cembratriene‐ol in the engineered S. cerevisiae BY4741 was increased by 2.22‐fold compared to the control by overexpressing ERG20 and cbts*Δp , respectively, using two promoters P GAP . Cembratriene‐ol was found to be successfully synthesized via the integrated expression of cbts*Δp , ggpps and ERG20 on the genome of S. cerevisiae BY4741. The titer of cembratriene‐ol in S. cerevisiae S25 was further increased by 1.80‐fold compared to the control via dynamic control of the squalene synthase gene ERG9 . Overexpression of the genes cbts*Δp and ggpps using pY26‐GPD‐TEF in S. cerevisiae S25 with their integration expression increased the titer of cembratriene‐ol by 26.1‐fold compared to S. cerevisiae S25. The titer of cembratriene‐ol was significantly enhanced by mitochondrial compartmentalized expression of cbts*Δp and ggpps , which was 76.3‐fold higher than that of the initial strain constructed. It was indicated that the systematic optimization has great potential in facilitating high‐level production of cembratriene‐ol production in S. cerevisiae .