Combined metabolic engineering and lipid droplets degradation to increase vitamin A production in Saccharomyces cerevisiae

作者：J.-B. Lin, Xiao Bu, Yibin Lan, Chang‐Qing Duan, Guoliang Yan · 发表于：Microbial Cell Factories · 年份：2024 · DOI：10.1186/s12934-024-02596-7 · 被引用次数：13 · 研究领域：Microbial Metabolic Engineering and Bioproduction、Algal biology and biofuel production、Lipid metabolism and biosynthesis

BACKGROUND: In microbial cell factories, substrate accessibility to enzyme is a key factor affecting the biosynthesis of natural products. As a robust chassis cells for biofuels and bioproducts, Saccharomyces cerevisiae also encounters the challenge since different enzymes and precursors are typically compartmentalized in different organelles. Such spatial separation could largely limit the efficiency of enzymatic reactions. In this study, the production of the hydrophobic product (vitamin A) was highly improved by metabolic engineering combined with degrading lipid droplets (the primary organelle storing β-carotene) to achieve efficient contact between β-carotene and 15, 15'-β-carotene monooxygenases in Saccharomyces cerevisiae. RESULTS: To efficiently produce vitamin A in Saccharomyces cerevisiae, ten 15, 15'-β-carotene monooxygenases (BCMOs) were firstly evaluated. The strain carrying marine bacterium 66A03 (Mb. BCMO) achieved the highest vitamin A titer. Co-adding 10% dodecane and 1% dibutylhydroxytoluene increased vitamin A titer to 19.03 mg/L in two-phase fermentation. Since most β-carotene is stored in LDs while BCMO is located in the cytosol, we developed a strategy to release β-carotene from LDs to better contact with BCMO. By overexpressing TGL3 and TGL4 using an ion-responsive promoter after high accumulation of β-carotene in LDs, LDs were sequentially degraded, which dramatically improved vitamin A production. Finally, by overexpressing tHMG1, ERG20, and CrtI and ...