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Abstract <p>Coenzyme Q10 biosynthesis in&#xa0;<i>Escherichia coli</i>&#xa0;is constrained by kinetic mismatches between precursor synthesis and methylation, alongside bioenergetic uncoupling. We implemented an optogenetic phase-control strategy integrating dynamic light induction, ribosome binding site (RBS) engineering, and real-time membrane potential (ΔΨ) feedback. Temporal coordination of 1-deoxy-D-xylulose-5-phosphate synthase (DXS) and&#xa0;UbiG methyltransferase (UbiG)&#xa0;via a 6-h phase delay reduced methylglyoxal shunt flux by 41 ± 3% (<i>p</i> &lt; 0.01) through enhanced precursor channeling. Membrane hyperpolarization to − 90 ± 2&#xa0;mV&#xa0;(relative to − 70&#xa0;mV in controls) triggered voltage-gated UbiG membrane localization (62 ± 3%) and ATP-driven S-adenosylmethionine regeneration, increasing methylation efficiency 2.3-fold. Multivariate modeling identified ΔΨ and acetate as critical control parameters, enabling optimized fermentation (dissolved oxygen (DO) 15–20%, pH 6.7–6.9). The engineered strain achieved 0.63 ± 0.07&#xa0;g/L CoQ10 in 5-L bioreactors—a 4.3-fold improvement over the static control strain (0.15 ± 0.02&#xa0;g/L)—with 82.5% carbon efficiency and 25.8% glycerol-to-product yield. This work establishes bioenergetically coupled temporal control as a scalable paradigm for membrane-bound isoprenoid biomanufacturing.</p> Key points <p>• <i>Phase-driven enzyme synchronization via optogenetics resolves kinetic mismatch.</i></p> <p>• <i>Membrane hyperpolarization gates enzyme localization and ATP regeneration.</i></p> <p>• <i>Model-integrated bioenergetic-process control enhances CoQ10 production efficiency.</i></p>

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Phase-driven rewiring in Escherichia coli enhances coenzyme Q10 biosynthesis via temporal and energetic coordination

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年11月24日

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  • Hao Li,
  • Yongjun Wu,
  • Dongfang Qin,
  • Jun Xie,
  • Wei Huang,
  • Ronghua Chen,
  • Hongmei Liu,
  • Ying Wang,
  • Dandan Zhao

摘要

Abstract

Coenzyme Q10 biosynthesis in Escherichia coli is constrained by kinetic mismatches between precursor synthesis and methylation, alongside bioenergetic uncoupling. We implemented an optogenetic phase-control strategy integrating dynamic light induction, ribosome binding site (RBS) engineering, and real-time membrane potential (ΔΨ) feedback. Temporal coordination of 1-deoxy-D-xylulose-5-phosphate synthase (DXS) and UbiG methyltransferase (UbiG) via a 6-h phase delay reduced methylglyoxal shunt flux by 41 ± 3% (p < 0.01) through enhanced precursor channeling. Membrane hyperpolarization to − 90 ± 2 mV (relative to − 70 mV in controls) triggered voltage-gated UbiG membrane localization (62 ± 3%) and ATP-driven S-adenosylmethionine regeneration, increasing methylation efficiency 2.3-fold. Multivariate modeling identified ΔΨ and acetate as critical control parameters, enabling optimized fermentation (dissolved oxygen (DO) 15–20%, pH 6.7–6.9). The engineered strain achieved 0.63 ± 0.07 g/L CoQ10 in 5-L bioreactors—a 4.3-fold improvement over the static control strain (0.15 ± 0.02 g/L)—with 82.5% carbon efficiency and 25.8% glycerol-to-product yield. This work establishes bioenergetically coupled temporal control as a scalable paradigm for membrane-bound isoprenoid biomanufacturing.

Key points

• Phase-driven enzyme synchronization via optogenetics resolves kinetic mismatch.

• Membrane hyperpolarization gates enzyme localization and ATP regeneration.

• Model-integrated bioenergetic-process control enhances CoQ10 production efficiency.