Biological CO₂ activation and conversion to high-value ethanol are a feasible and green strategy to close the carbon cycle. However, naturally evolved CO₂ utilization pathways involve carbon loss or ATP consumption. Herein, we report a complete anabolic pathway for direct conversion of CO₂ to ethanol by constructing and assembling three functional modules including CO₂ activation, formaldehyde → acetyl-CoA, and ethanol synthesis in a carbon-conserved and ATP-independent system. These artificial ethanol metabolic pathway fluxes were strengthened by screening efficient key enzymes for CO₂ activation, promoting formaldehyde assimilation, and employing a two-step reaction. ¹³C-labeled CO₂ demonstrated the feasibility of the pathway converting CO₂ into ethanol in vitro by detecting the carbon flow. With this anabolic pathway, we obtained an ethanol concentration of 0.37 mM at a conversion rate of 4.33 nmol CO₂ min⁻¹ mg⁻¹ enzyme and 0.5 mM R5P supply. This modularization strategy provides a new avenue in the construction of artificial metabolic pathways for ethanol synthesis from CO₂.

中文翻译：

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将二氧化碳直接转化为乙醇的无细胞人工合成代谢途径

生物CO ₂活化并转化为高价值乙醇是闭合碳循环的可行且绿色的策略。然而，自然进化的CO ₂利用途径涉及碳损失或ATP消耗。在此，我们通过在碳保守且不依赖于ATP的系统中构建和组装三个功能模块（包括CO 2 活化、甲醛→乙酰辅酶A和乙醇合成），报告了将CO 2 直接转化为_乙醇的完整合成代谢途径。_{通过筛选有效的CO 2}活化关键酶、促进甲醛同化以及采用两步反应，增强了这些人工乙醇代谢途径通量。¹³ C标记的CO ₂通过检测碳流证明了体外将CO ₂转化为乙醇的途径的可行性。_{通过这种合成代谢途径，我们以 4.33 nmol CO 2} min ^-1 mg ^-1酶和 0.5 mM R5P 供应的转化率获得了 0.37 mM 的乙醇浓度。这种模块化策略为构建从CO ₂合成乙醇的人工代谢途径提供了新途径。