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Background <p>3-Hydroxypropionic acid (3-HP) is a promising C3 platform chemical with wide industrial applications. However, its microbial production remains limited by insufficient intracellular malonyl-CoA availability and metabolic imbalance.</p> Result <p>In this study, we systematically engineered <i>Escherichia coli</i> for enhanced 3-HP biosynthesis. The malonate assimilation genes (<i>matB</i>, <i>smatPQM</i>) and 3-HP biosynthesis gene (<i>mcr</i>) were chromosomally integrated using CRISPR/Cas9, resulting in a plasmid-free, antibiotic-free strain (WYY04) that produced 21.97 mM 3-HP, 0.51-fold higher than the plasmid-based system. Further improvement was achieved by CRISPRi-mediated repression of fatty acid biosynthesis genes (<i>fabD</i>, <i>fabF</i>), increasing 3-HP titer by 66%. Introduction of a malonyl-CoA-responsive FapR/fapO biosensor enabled dynamic regulation of <i>mcr</i> expression, enhancing 3-HP production by 59%. Through all these above engineering, the 3-HP production of the strain WYY19 increased by 2.29 times compared to that of the plasmid-expressing system. Under optimized fermentation conditions, the final engineered strain WYY19 produced 42.22&#xa0;g/L 3-HP with the specific productivity of 0.69&#xa0;g/g and 0.46&#xa0;g/L/h from glucose and malonate in fed-batch bioreactor.</p> Conclusions <p>This study demonstrates a robust, genetically stable, and scalable microbial platform for 3-HP biosynthesis.</p>

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Systematic engineering of Escherichia coli for biosynthesis of 3-hydroxypropionic acid from glucose and malonate

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2026年3月24日

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  • Yingying Wang,
  • Meihui Cao,
  • Mei Hu,
  • Hengrui Xu,
  • Yafei Du,
  • Changxing Sun,
  • Lingfei Kong,
  • Yu Luo,
  • Xiaoyu Liu,
  • Jianming Yang,
  • Bo Liang

摘要

Background

3-Hydroxypropionic acid (3-HP) is a promising C3 platform chemical with wide industrial applications. However, its microbial production remains limited by insufficient intracellular malonyl-CoA availability and metabolic imbalance.

Result

In this study, we systematically engineered Escherichia coli for enhanced 3-HP biosynthesis. The malonate assimilation genes (matB, smatPQM) and 3-HP biosynthesis gene (mcr) were chromosomally integrated using CRISPR/Cas9, resulting in a plasmid-free, antibiotic-free strain (WYY04) that produced 21.97 mM 3-HP, 0.51-fold higher than the plasmid-based system. Further improvement was achieved by CRISPRi-mediated repression of fatty acid biosynthesis genes (fabD, fabF), increasing 3-HP titer by 66%. Introduction of a malonyl-CoA-responsive FapR/fapO biosensor enabled dynamic regulation of mcr expression, enhancing 3-HP production by 59%. Through all these above engineering, the 3-HP production of the strain WYY19 increased by 2.29 times compared to that of the plasmid-expressing system. Under optimized fermentation conditions, the final engineered strain WYY19 produced 42.22 g/L 3-HP with the specific productivity of 0.69 g/g and 0.46 g/L/h from glucose and malonate in fed-batch bioreactor.

Conclusions

This study demonstrates a robust, genetically stable, and scalable microbial platform for 3-HP biosynthesis.