Electrocatalytic CO<sub>2</sub> Reduction Coupled with Metabolic Engineering of <italic style="font-style: italic">Yarrowia lipolytica</italic> for Efficient Isoprene Biosynthesis

LAN Yunlong; LI Song; ZHANG Wei; CHANG Yunyun; LIU Yanhui

doi:10.12211/2096-8280.2025-046

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Electrocatalytic CO₂ Reduction Coupled with Metabolic Engineering of Yarrowia lipolytica for Efficient Isoprene Biosynthesis

更新时间：2025-09-17

- Electrocatalytic CO₂ Reduction Coupled with Metabolic Engineering of Yarrowia lipolytica for Efficient Isoprene Biosynthesis
- Synthetic Biology Journal Pages: 1-15(2025)
- 作者机构：
  
  1.绿色生物制造全国重点实验室，北京 100029
  2.北京化工大学国家能源生物炼制研发中心，北京 100029
  3.北京化工大学生命科学与技术学院，北京 100029
- 作者简介：
- 基金信息：
- DOI：10.12211/2096-8280.2025-046
  CLC： Q939.9;Q789
- Received：19 May 2025，
  
  Revised：2025-09-15，
  
  Online First：17 September 2025，
- 稿件说明：
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兰云龙，李嵩，张伟，常允蕴，刘艳辉. 电催化还原CO₂耦合代谢工程改造解脂耶氏酵母实现高效异戊二烯生物合成［J］. 合成生物学， 2025， 6. DOI： 10.12211/2096-8280.2025-046

LAN Yunlong， LI Song， ZHANG Wei， CHANG Yunyun， LIU Yanhui. Electrocatalytic CO₂ Reduction Coupled with Metabolic Engineering of Yarrowia lipolytica for Efficient Isoprene Biosynthesis［J］. Synthetic Biology Journal， 2025， 6. DOI： 10.12211/2096-8280.2025-046
兰云龙，李嵩，张伟，常允蕴，刘艳辉. 电催化还原CO₂耦合代谢工程改造解脂耶氏酵母实现高效异戊二烯生物合成［J］. 合成生物学， 2025， 6. DOI： 10.12211/2096-8280.2025-046 DOI：

LAN Yunlong， LI Song， ZHANG Wei， CHANG Yunyun， LIU Yanhui. Electrocatalytic CO₂ Reduction Coupled with Metabolic Engineering of Yarrowia lipolytica for Efficient Isoprene Biosynthesis［J］. Synthetic Biology Journal， 2025， 6. DOI： 10.12211/2096-8280.2025-046 DOI：

摘要

生物合成技术因其绿色可持续特性，成为替代传统石化工艺的重要方向。本研究通过电催化还原CO₂与代谢工程改造解脂耶氏酵母相结合的策略，实现了高效异戊二烯的生物合成。首先，在解脂耶氏酵母中异源表达不同来源的异戊二烯合酶，筛选出葛根来源的异戊二烯合酶，异戊二烯产量为560 μg/L。随后，通过强化甲羟戊酸途径关键基因

erg10、erg12、erg13、erg8、erg19、thmgr、idi

及下调竞争基因

erg20

，构建工程菌株Misps13，摇瓶发酵异戊二烯滴度提高至12.23 mg/L。进一步整合电催化-微生物耦合系统，利用自组装纳米材料L-Bi-sh-H₂O和L-Cu-sh-3将CO₂高效转化为甲酸和乙酸，其法拉第效率分别为88.67%和50.14%。在Misps13中引入ACS与FDH模块，使工程菌Misps1315能够利用电催化合成的甲酸和乙酸，增强胞内乙酰辅酶A和NAD（P）H供应。通过响应曲面法优化外源甲酸和乙酸添加量，其中甲酸2.8 g/L、乙酸6.5 g/L，最终实现异戊二烯滴度32.14 mg/L，较基础菌株提升2.6倍。本研究为生物-电化学协同制造高值化合物提供了新范式。

Abstract

Isoprene is a valuable platform chemical essential for manufacturing synthetic rubber

elastomers

and specialty materials

yet its conventional production depends heavily on petroleum resources with significant carbon emissions. Here

we present an innovative bio-electrocatalytic process that synergistically combines CO

electroreduction with metabolically engineered

Yarrowia lipolytica

for sustainable isoprene production. We began by screening several plant-derived isoprene synthases and identified the enzyme from

Pueraria lobata

(Isps-Pu) as the most effective in this yeast host

yielding initial production of 560 μg/L. Subsequent rational metabolic engineering involved systematic overexpression of the mevalonate pathway genes

including

erg10

erg12

erg13

erg8

erg19

thmgr

and

idi

via iterative genomic integration. Crucially

competitive metabolic flux was reduced by replacing the native promoter of

erg20

with the weak promoter P

KI1

resulting in a significantly improved isoprene titer of 12.23 mg/L.To establish an efficient electrocatalytic module

we systematically developed and characterized multiple nanomaterials for CO₂ reduction. For formate production

bismuth-based electrodes (L-Bi-sh-C and L-Bi-sh-H₂O) were synthesized through distinct self-assembly and thermal treatment pathways. Comparative analysis revealed that L-Bi-sh-H

prepared via hydrothermal treatment

demonstrated superior performance with a remarkable Faradaic efficiency of 88.67% for formate at -1.8 V versus Ag/AgCl. For acetate production

we engineered a series of copper-based catalysts (L-Cu-sh-3

L-Cu-sh-3-H₂O

L-Cu-sh-EtOH

and L-Cu-sh-EtOH-H

O) using different solvent systems and processing methods. Among these

L-Cu-sh-3 electrode synthesized in acetonitrile/methanol/tetrahydrofuran solvent mixture exhibited optimal performance

achieving 50.14% Faradaic efficiency for acetate at -0.8 V.The integration of these electrocatalytic components with biological conversion was achieved by introducing

Saccharomyces cerevisiae

-derived acetyl-CoA synthase and formate dehydrogenase into the engineered yeast

enabling efficient assimilation of electro-generated formate and acetate to enhance intracellular acetyl-CoA and NADPH pools. Through response surface methodology optimization

we determined optimal concentrations of formate (2.8 g/L) and acetate (6.5 g/L)

leading to a final isoprene titer of 32.14 mg/L—representing a 2.6-fold enhancement over the baseline strain. This integrated approach not only demonstrates a carbon-negative strategy for isoprene biosynthesis but also establishes a versatile platform for producing acetyl-CoA-derived chemicals from CO₂.

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Keywords

references

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⁰

Electrocatalytic CO2 Reduction Coupled with Metabolic Engineering of Yarrowia lipolytica for Efficient Isoprene Biosynthesis

DOI：10.12211/2096-8280.2025-046

摘要

Abstract

关键词

Keywords

references

Electrocatalytic CO₂ Reduction Coupled with Metabolic Engineering of Yarrowia lipolytica for Efficient Isoprene Biosynthesis