Scholarlink

Background <p>Methanol is a promising one-carbon (C1) feedstock for sustainable bioproduction, and its mixotrophic co-utilization with other substrates can improve product formation. However, mixotrophy often leads to sequential substrate utilization that delays methanol assimilation, and the regulatory basis underlying this phenotype remains unclear. In this study, we aimed to elucidate the regulatory mechanism governing methanol utilization in a methylotrophic acetogen and to determine whether rewiring this system could improve methanol co-utilization.</p> Results <p>Here, we identify a dual-layer regulatory circuit centered on P<sub>mtaR</sub>, the promoter driving the <i>mta</i> operon in <i>Eubacterium limosum</i>, as the key regulatory locus where methanol-responsive activation and carbon catabolite repression are integrated to govern the onset of methanol utilization. We show that robust P<sub>mtaR</sub> activation requires the AraC-type regulator MtaR along with an upstream activation region within the promoter, whereas this activation is counteracted by a catabolite-responsive element (<i>cre</i>) embedded in P<sub>mtaR</sub>, consistent with CcpA-mediated repression. This dual-layer regulatory architecture explains the delayed induction of the <i>mta</i> operon and the sequential utilization of glucose and methanol in <i>E. limosum</i>. Rewiring <i>mta</i> expression with a <i>cre</i>-free methanol-responsive promoter relieved repression enabled improved glucose–methanol co-utilization with enhanced methanol assimilation during glucose consumption. This achieved up to 3-fold increases in growth, substrate uptake, and product formation rates, accompanied by a metabolic shift towards butyrate production.</p> Conclusions <p>This study reveals a dual regulatory mechanism governing methanol utilization in <i>E. limosum</i> by integrating local methanol-responsive activation with global carbon catabolite repression. This mechanism explains sequential substrate utilization during glucose–methanol mixotrophy and provides a practical engineering strategy to improve methanol co-utilization and product formation in acetogenic bioprocesses.</p>

错误：搜索内容不能为空，请输入英文关键词
错误：关键词超出字数限制，请精简
高级检索

Rewiring a methanol-responsive regulatory system improves glucose–methanol co-utilization in Eubacterium limosum

  • 期刊论文

  • 出版模式：

    开放获取

  • 发表日期：

    2026年7月3日

查看全文
  • Jiyun Bae,
  • Hyunwoo Jung,
  • Seulgi Kang,
  • Chanho Park,
  • Donghwi Lee,
  • Sangrak Jin,
  • Byung-Kwan Cho

摘要

Background

Methanol is a promising one-carbon (C1) feedstock for sustainable bioproduction, and its mixotrophic co-utilization with other substrates can improve product formation. However, mixotrophy often leads to sequential substrate utilization that delays methanol assimilation, and the regulatory basis underlying this phenotype remains unclear. In this study, we aimed to elucidate the regulatory mechanism governing methanol utilization in a methylotrophic acetogen and to determine whether rewiring this system could improve methanol co-utilization.

Results

Here, we identify a dual-layer regulatory circuit centered on PmtaR, the promoter driving the mta operon in Eubacterium limosum, as the key regulatory locus where methanol-responsive activation and carbon catabolite repression are integrated to govern the onset of methanol utilization. We show that robust PmtaR activation requires the AraC-type regulator MtaR along with an upstream activation region within the promoter, whereas this activation is counteracted by a catabolite-responsive element (cre) embedded in PmtaR, consistent with CcpA-mediated repression. This dual-layer regulatory architecture explains the delayed induction of the mta operon and the sequential utilization of glucose and methanol in E. limosum. Rewiring mta expression with a cre-free methanol-responsive promoter relieved repression enabled improved glucose–methanol co-utilization with enhanced methanol assimilation during glucose consumption. This achieved up to 3-fold increases in growth, substrate uptake, and product formation rates, accompanied by a metabolic shift towards butyrate production.

Conclusions

This study reveals a dual regulatory mechanism governing methanol utilization in E. limosum by integrating local methanol-responsive activation with global carbon catabolite repression. This mechanism explains sequential substrate utilization during glucose–methanol mixotrophy and provides a practical engineering strategy to improve methanol co-utilization and product formation in acetogenic bioprocesses.