Adaptive Evolution and Metabolic Engineering Boost Lycopene Production in Saccharomyces cerevisiae via Enhanced Precursors Supply and Utilization

Lycopene is a red carotenoid with remarkable antioxidant activity, which has been widely used in food, cosmetics, medicine, and other industries. Production of lycopene in Saccharomyces cerevisiae provides an economic and sustainable means. Many efforts have been done in recent years, but the titer of lycopene seems to reach a ceiling. Enhancing the supply and utilization of farnesyl diphosphate (FPP) is generally regarded as an efficient strategy for terpenoid production. Herein, an integrated strategy by means of atmospheric and room-temperature plasma (ARTP) mutagenesis combined with H2O2-induced adaptive laboratory evolution (ALE) was proposed to improve the supply of upstream metabolic flux toward FPP. Enhancing the expression of CrtE and introducing an engineered CrtI mutant (Y160F&N576S) increased the utilization of FPP toward lycopene. Consequently, the titer of lycopene in the strain harboring the Ura3 marker was increased by 60% to 703 mg/L (89.3 mg/g DCW) at the shake-flask level. Eventually, the highest reported titer of 8.15 g/L of lycopene in S. cerevisiae was achieved in a 7 L bioreactor. The study highlights an effective strategy that the synergistic complementarity of metabolic engineering and adaptive evolution facilitates natural product synthesis.番茄红素（Lycopene）是一种红色类胡萝卜素（carotenoid），具备显著的抗氧化活性，已被广泛应用于食品、化妆品、医药等多个工业领域。利用酿酒酵母（Saccharomyces cerevisiae）生产番茄红素是一种经济且可持续的生产方式。尽管近年来已开展诸多研究工作，但番茄红素的发酵效价似乎已触及瓶颈。法尼基二磷酸（FPP）的供给与利用提升通常被视为萜类化合物（terpenoid）生产的高效策略。本研究提出一种整合策略：通过常压室温等离子体（ARTP）诱变结合过氧化氢（H₂O₂）诱导的适应性实验室进化（ALE），以增强上游代谢流向法尼基二磷酸（FPP）的通量。通过强化CrtE基因的表达，并引入工程化改造的CrtI突变体（Y160F&N576S），可提升法尼基二磷酸（FPP）向番茄红素的转化利用效率。最终，携带Ura3筛选标记的工程菌株在摇瓶水平下的番茄红素效价提升60%，达到703 mg/L（89.3 mg/g细胞干重（DCW））。在7 L生物反应器中，该菌株最终实现了目前酿酒酵母（Saccharomyces cerevisiae）中报道的最高番茄红素效价——8.15 g/L。本研究凸显了一项有效策略，即代谢工程与适应性进化的协同互补可有效促进天然产物的合成。