Joint International Research Laboratory of Agriculture and Agri-Product Safety, the Ministry of Education of China, Yangzhou University, Yangzhou, Jiangsu 225009, PR China.
College of Bioscience and Biotechnology, Yangzhou University, Yangzhou 225009, PR China.
J Agric Food Chem. 2021 Jan 27;69(3):1003-1010. doi: 10.1021/acs.jafc.0c06384. Epub 2021 Jan 11.
Linalool, as a fragrant monoterpene, is an important feedstock for food, pharmaceuticals, and cosmetics industries. Although our previous study had significantly increased linalool production by the directed evolution of linalool synthase and overexpression of the whole mevalonate pathway genes, the engineered yeast strain suffered from dramatically reduced biomass. Herein, a stress-free linalool-producing yeast cell factory was constructed by the combinational regulation of linalool synthase and farnesyl diphosphate synthase instead of multienzyme overexpression. First, the expression level of linalool synthase was successfully enhanced by introducing a N-terminal SKIK tag, which improved linalool production by 3.3-fold. Subsequently, the modular assembly of linalool synthase and dominant negative farnesyl diphosphate synthase via short peptide tags efficiently converted geranyl pyrophosphate to linalool. Additional downregulation of the native farnesyl diphosphate synthase led to the highest reported linalool production (80.9 mg/L) in yeast. This combinatorial modulation strategy may also be applied to the production of other high-value monoterpenes.
芳樟醇是一种重要的单萜类香料，是食品、制药和化妆品工业的重要原料。虽然我们之前的研究通过定向进化芳樟醇合酶和过表达整条甲羟戊酸途径的基因显著提高了芳樟醇的产量，但工程酵母菌株的生物量却大幅下降。在此，通过组合调控芳樟醇合酶和法呢基二磷酸合酶，而不是多酶过表达，构建了一个无应激的芳樟醇生产酵母细胞工厂。首先，通过引入 N 端 SKIK 标签成功提高了芳樟醇合酶的表达水平，使芳樟醇产量提高了 3.3 倍。随后，通过短肽标签将芳樟醇合酶和显性负性法呢基二磷酸合酶进行模块化组装，有效地将香叶基焦磷酸转化为芳樟醇。进一步下调内源性法呢基二磷酸合酶可导致酵母中报道的最高芳樟醇产量（80.9mg/L）。这种组合调控策略也可应用于其他高价值单萜类化合物的生产。
