Synthetic and Systems Biotechnology最新文献_第7页

Combination of enzyme engineering and quorum sensing system for efficient de novo biosynthesis of β-arbutin in Escherichia coli 结合酶工程和群体感应系统在大肠杆菌中高效从头合成β-熊果苷

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2026-03-01 Epub Date: 2025-08-22 DOI: 10.1016/j.synbio.2025.08.010

Zhuoyuan Liu , Dongming Liu , Lihao Deng , Kai LingHu , Xiaoyu Shan , Jingwen Zhou , Weizhu Zeng

β-Arbutin is a plant-derived glycoside with various biological activities, which has been applied in pharmaceuticals and whitening cosmetics. Uneven carbon flux distribution and the low catalytic efficiency of arbutin synthase limit the biosynthesis of β-arbutin in microorganisms. In addition, commonly used high-cost, or toxic, inducible expression agents are not practical to use in industrial production. In this study, a biosynthetic pathway for β-arbutin was constructed in Escherichia coli. The competition and degradation pathways were knocked out, and the key pathway enzymes were identified and overexpressed, to increase β-arbutin production. Furthermore, by combining modifications to the phosphoketolase pathway and the acetic acid pathway, the carbon flux was redirected to the shikimic acid pathway. The substrate access channel of the arbutin synthase active site was modified by protein engineering, resulting in the R241H mutant, which increased β-arbutin production by 16.7 %, compared with wild type (WT). Finally, the luxR-based quorum sensing (QS) system was used to regulate the allocation of metabolic fluxes between cell growth and product synthesis, achieving a self-induced regulation of E. coli, without requiring exogenous inducers. The final engineered strain produced 81.9 g/L of β-arbutin from a fed-batch fermentation in a 5 L fermenter and the yield from glucose reached 0.29 g/g. This study presents a systematic strategy that can provide a reference for the construction of β-arbutin microbial cell factories.

β-熊果苷是一种具有多种生物活性的植物源性糖苷，在医药、美白化妆品等领域有着广泛的应用。碳通量分布不均和熊果苷合酶催化效率低限制了微生物对β-熊果苷的生物合成。此外，常用的高成本或有毒的诱导表达剂不适合用于工业生产。本研究在大肠杆菌中构建了β-熊果苷的生物合成途径。敲除竞争和降解途径，鉴定并过表达关键途径酶，增加β-熊果苷的产量。此外，通过结合磷酸酮醇酶途径和乙酸途径的修饰，将碳通量重定向到莽草酸途径。通过蛋白工程修饰熊果苷合成酶活性位点的底物通路，得到的R241H突变体β-熊果苷产量较野生型（WT）提高16.7%。最后，利用基于luxr的群体感应（quorum sensing， QS）系统调节代谢通量在细胞生长和产物合成之间的分配，实现了大肠杆菌不需要外源诱导剂的自我诱导调节。最终工程菌株在5 L的发酵罐中分批补料发酵，β-熊果苷产量为81.9 g/L，葡萄糖产量为0.29 g/g。本研究为β-熊果苷微生物细胞工厂的建设提供了系统的策略参考。

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引用次数: 0

Putative membrane protein Cgl2168 and its variant enhance thermotolerance in Corynebacterium glutamicum 推测膜蛋白Cgl2168及其变体增强谷氨酸棒状杆菌的耐热性

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2026-03-01 Epub Date: 2025-09-25 DOI: 10.1016/j.synbio.2025.09.018

Ning Gao , Xiaomeng Ni , Jiao Liu , Jiuzhou Chen , Yu Wang , Ping Zheng , Jibin Sun

Corynebacterium glutamicum ATCC 13032 is widely used as the chassis strain for industrial production of various amino acids, but its thermotolerance limits productivity under high temperatures. Comparative genomic analysis in this study identified cgl2168 as a key determinant of enhanced thermotolerance in a laboratory strain (13032-JN) compared to another stock (13032-TJ). A guanine at Residue 270 (G270) deletion in cgl2168 (cgl2168^Δ270G) in 13032-JN significantly improved growth at 40 °C, while reverting this mutation or deleting cgl2168 impaired thermotolerance. Structural analysis revealed the deletion extends the C-terminus with an additional α-helix. Transcriptomic profiling showed cgl2168^Δ270G increased gene expressions involved in oxidative phosphorylation (e.g., atpABCDEFGH), nitrate metabolism (narIJHK), some heat shock protein genes (e.g., dnaK and groES) and deletion of cgl2168 resulted in upregulating sulfur metabolism (sulfonate transporter protein genes ssuBCD, sulfur-containing amino acid cys and met operons) and central carbon metabolism (gltA, pyc), while downregulating gluconeogenesis (pck) and fermentation (ldh) genes. These changes enhanced ATP synthesis, carbon utilization and oxidative tolerance. Predicted interactors of Cgl2168 linked it to respiratory chain function, stress responses, and cell envelope biogenesis. Thus, Cgl2168 and its variant Cgl2168^Δ270G enhance thermotolerance by coordinating energy production, carbon metabolism, and stress adaptation, aiding industrial applications.

谷氨棒状杆菌ATCC 13032被广泛用作各种氨基酸工业生产的基础菌株，但其耐热性限制了高温下的生产能力。本研究的比较基因组分析发现，cgl2168是实验室菌株（13032-JN）与另一个品系（13032-TJ）相比耐热性增强的关键决定因素。13032-JN中cgl2168 （cgl2168Δ270G）中残基270处的鸟嘌呤（G270）缺失显著改善了40°C下的生长，同时恢复该突变或删除cgl2168损伤了耐热性。结构分析显示，该缺失扩展了c端，增加了一个α-螺旋。转录组学分析显示cgl2168Δ270G参与氧化磷酸化（如atpABCDEFGH）、硝酸盐代谢（narIJHK）、一些热休克蛋白基因（如dnaK和groES）的基因表达增加，cgl2168的缺失导致硫代谢（磺酸转运蛋白基因ssuBCD、含硫氨基酸cys和met操纵子）和中心碳代谢（gltA、pyc）基因表达上调，糖异生（pck）和发酵（ldh）基因表达下调。这些变化增强了ATP合成、碳利用和氧化耐受性。预测的Cgl2168相互作用物将其与呼吸链功能、应激反应和细胞包膜生物发生联系起来。因此，Cgl2168及其变体Cgl2168Δ270G通过协调能量产生、碳代谢和逆境适应来增强热耐受性，有助于工业应用。

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引用次数: 0

Competitive binding of TeXlnR and TeAmyR regulates cellulase production in Talaromyces endophyticus TeXlnR和TeAmyR的竞争结合调控了Talaromyces内生纤维素酶的产生

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2026-03-01 Epub Date: 2025-11-03 DOI: 10.1016/j.synbio.2025.10.010

Yixin Sun , Qi Sun , Tiantian Gao , Shumin Ren , Xiaomei Hu

Efficient cellulase production is essential for its application in green bioprocessing, yet the transcriptional regulation of cellulase biosynthesis in Talaromyces sp. remains largely uncharacterized. In this study, we elucidate the regulatory functions of TeXlnR and TeAmyR in Talaromyces endophyticus. Our results demonstrate that TeXlnR, as a potent transcriptional activator, enhances enzyme production by over 33 %. RNA-seq analysis shows that TeXlnR overexpression upregulates 33 cellulase genes and 11 hemicellulase genes, while concurrently repressing the expression of the transcription factor TeAmyR. Further investigation identified TeAmyR as a repressor of cellulase production in T. endophyticus. EMSA and yeast one-hybrid analyses confirmed the specific competitive binding of these two transcription factors to the promoter regions of cellulase genes eg5B and eg12A. TeXlnR both directly activates target gene transcription and indirectly promotes cellulase synthesis by downregulating TeAmyR expression to reduce its competitive binding to promoters. These findings provide direct evidence for competitive regulation by transcription factors in Talaromyces sp. and establish a theoretical foundation for the rational engineering of industrial strains with improved cellulase production.

高效的纤维素酶生产对于其在绿色生物加工中的应用至关重要，然而Talaromyces sp.纤维素酶生物合成的转录调控在很大程度上仍未被表征。在本研究中，我们阐明了TeXlnR和TeAmyR在藻内生中的调控作用。我们的研究结果表明，TeXlnR作为一种有效的转录激活剂，可以使酶的产量提高33%以上。RNA-seq分析显示，TeXlnR过表达上调33个纤维素酶基因和11个半纤维素酶基因，同时抑制转录因子TeAmyR的表达。进一步的研究发现，TeAmyR是内生葡萄球菌纤维素酶生产的抑制因子。EMSA和酵母单杂交分析证实了这两个转录因子与纤维素酶基因eg5B和eg12A的启动子区特异性竞争结合。TeXlnR既可以直接激活靶基因转录，又可以通过下调TeAmyR的表达，减少其与启动子的竞争性结合，间接促进纤维素酶的合成。这些发现为Talaromyces sp.受转录因子竞争调控提供了直接证据，并为提高纤维素酶产量的工业菌株的合理工程设计奠定了理论基础。

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引用次数: 0

Harnessing prokaryotic amino acid transporters for metabolic engineering: mechanisms and biotechnological applications 利用原核氨基酸转运体进行代谢工程：机制和生物技术应用

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2026-03-01 Epub Date: 2025-10-22 DOI: 10.1016/j.synbio.2025.10.007

Junyao Huang , Wenming Wu , Zhicheng Fu , Chunyu Chao , Chongde Lai , Xiaoyu Wu , Hui Yang , Xiaohe Chu , Bangce Ye , Bin Zhang

As fundamental bioactive molecules and building blocks of cellular structures, amino acids play a crucial role in sustaining life, reflected in a global market valued at tens of billions of dollars. Recently, microbial fermentation has become a leading alternative to chemical synthesis and enzymatic catalysis for amino acid production, driving the need for robust strains. Efficient transport of amino acids out of the cell is essential for strain development, as it mitigates intracellular accumulation, cytotoxicity, and feedback inhibition, thereby improving fermentation efficiency. This review summarizes the physiological characteristics of amino acid transporters and their applications in the metabolic engineering of high-performance microbial strains. In particular, it provides a comprehensive overview of how amino acid transport systems are applied to develop amino acid-producing strains by enhancing substrate uptake, increasing precursor supply, directly promoting amino acid transportation, and reducing flux through competing metabolic pathways. Furthermore, this review explores future challenges and prospects in the field, offering theoretical guidance for engineering robust microbial platforms capable of producing not only amino acids but also high-value bioproducts.

作为基本的生物活性分子和细胞结构的组成部分，氨基酸在维持生命中起着至关重要的作用，这反映在价值数百亿美元的全球市场上。最近，微生物发酵已成为化学合成和酶催化氨基酸生产的主要替代方法，推动了对健壮菌株的需求。氨基酸在细胞外的有效运输对菌株的发育至关重要，因为它减轻了细胞内的积累、细胞毒性和反馈抑制，从而提高了发酵效率。本文综述了氨基酸转运体的生理特性及其在高效微生物菌株代谢工程中的应用。特别是，它提供了氨基酸运输系统如何通过增强底物摄取、增加前体供应、直接促进氨基酸运输和通过竞争代谢途径减少通量来应用于氨基酸生产菌株的全面概述。此外，本文还探讨了该领域未来的挑战和前景，为构建既能生产氨基酸又能生产高价值生物制品的强大微生物平台提供理论指导。

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引用次数: 0

Efficient synthesis of l-DOPA in Escherichia coli via cofactor and enzyme engineering 利用辅助因子和酶工程在大肠杆菌中高效合成左旋多巴

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2026-03-01 Epub Date: 2025-09-12 DOI: 10.1016/j.synbio.2025.09.011

Lihao Deng , Jurong Ping , Zhuoyuan Liu , Kai Linghu , Heng Zhang , Xiaoyu Shan , Weizhu Zeng , Jianghua Li , Jingwen Zhou

The global incidence of Parkinson's disease continues to rise. Levodopa (l-DOPA) is the core therapeutic drug, and efficient and sustainable production methods are needed. However, the complex metabolic pathways and the low catalytic efficiency of enzymes limit biosynthesis of l-DOPA in microorganisms. To address this issue, this study significantly enhanced the production efficiency of l-DOPA through a multi-dimensional, integrated metabolic and enzyme engineering approach. Firstly, the de novo synthesis pathway for l-DOPA was established through optimization of the promoter, ribosome-binding site (RBS), plasmid copy number, and tighly accurately regulating the expression level of key enzymes. Secondly, combined with metabonomic analysis, carbon metabolic flow was diverted, increasing the l-DOPA titer by 36.7 %. Glucose dehydrogenase (BmgdH) and gluconate kinase (gntK) were introduced to construct a cofactor regeneration system, which synergistically enhanced the supply of NADH and FADH₂, increasing the l-DOPA conversion rate by 18 %. Next, the substrate tunnel of 4-hydroxyphenylacetic acid-3-monooxygenase subunit B (HpaB) was subjected to rational design, and mutant T292A significantly expanded the substrate channel, improved catalytic efficiency, and decreased l-tyrosine by 87 %. Finally, through the process optimization in a 5 L bioreactor (involving phased pH control and induction timing adjustment) achieved an l-DOPA titer of 60.73 g/L, the highest reported to date for de novo microbial synthesis. This research offers a novel approach for industrial biosynthesis of l-DOPA, and broadens engineering concepts for efficient synthesis of aromatic compounds.

帕金森氏症的全球发病率持续上升。左旋多巴（l-DOPA）是核心治疗药物，需要高效、可持续的生产方法。然而，复杂的代谢途径和酶的低催化效率限制了微生物对左旋多巴的生物合成。为了解决这一问题，本研究通过多维、综合的代谢和酶工程方法，显著提高了左旋多巴的生产效率。首先，通过优化启动子、核糖体结合位点（RBS）、质粒拷贝数，严格准确调控关键酶的表达水平，建立l-DOPA从头合成途径。其次，结合代谢组学分析，改变了碳代谢流量，使左旋多巴滴度提高了36.7%。引入葡萄糖脱氢酶（BmgdH）和葡萄糖酸激酶（gntK）构建辅助因子再生体系，协同增强NADH和FADH2的供应，使l-DOPA转化率提高18%。接下来，对4-羟基苯基乙酸-3-单加氧酶亚基B （HpaB）的底物通道进行合理设计，突变体T292A显著扩大了底物通道，提高了催化效率，使l-酪氨酸降低了87%。最后，通过在5l生物反应器中进行工艺优化（包括分阶段pH控制和诱导时间调节），获得了60.73 g/L的L - dopa滴度，这是迄今为止报道的微生物从头合成的最高滴度。本研究为左旋多巴的工业生物合成提供了新的途径，拓宽了芳香族化合物高效合成的工程概念。

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引用次数: 0

Identification of a novel β-1,4-galactosyltransferase from Haemophilus parainfluenzae for efficient biosynthesis of lacto-N-neotetraose in Escherichia coli 副流感嗜血杆菌β-1,4-半乳糖转移酶在大肠杆菌中高效合成乳酸-n -新四糖的鉴定

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2026-03-01 Epub Date: 2025-09-05 DOI: 10.1016/j.synbio.2025.09.006

Dan Liu , Chenhui Ying , Yingqi Ping , Shanquan Liang , Qiaojuan Yan , Zhengqiang Jiang

As an important member of human milk oligosaccharides (HMOs), lacto-N-neotetraose (LNnT) has been recognized as a promising HMO with many physiological activities for infants. β-1,4-Galactosyltransferases play a vital role in the final step of LNnT biosynthesis in microbial cell factory. Herein, we identified a novel β-1,4-galactosyltransferase (Hpa-GalT) from Haemophilus parainfluenzae for the efficient biosynthesis of LNnT in E. coli BL21 star(DE3). After overexpression of the key genes in the synthetic pathway, the titer of LNnT reached 1.86 g/L in shake flask fermentation. Moreover, a variant A186P/E146Q of Hpa-GalT was obtained by molecular modification, resulting in an increase of the LNnT titer to 2.15 g/L. Subsequently, a series of genes related to competitive pathways were deleted, including lacZ, ugd, wcaJ, and wecB, the engineered strain produced a LNnT titer of 2.23 g/L. After optimization of LNnT transport along with LNT II utilization, the engineered strain AH31 produced 2.93 g/L of LNnT in shake flask fermentation. Furthermore, it produced a maximum titer of 32.6 g/L of total LNnT and 27.1 g/L of extracellular LNnT in a 5-L fermentor with the productivity for 0.52 g/L/h. Overall, this study provides a novel β-1,4-galactosyltransferase and lays a foundation for the efficient biosynthesis of LNnT.

乳清-n -新四糖（LNnT）作为人乳寡糖（HMOs）的重要成员，具有多种婴儿生理活性，是一种很有发展前景的低聚糖。在微生物细胞工厂中，β-1,4-半乳糖转移酶在LNnT生物合成的最后一步起着至关重要的作用。在此，我们从副流感嗜血杆菌中鉴定出一种新的β-1,4-半乳糖转移酶（Hpa-GalT），用于大肠杆菌BL21星（DE3）中高效的LNnT生物合成。合成途径关键基因过表达后，摇瓶发酵LNnT滴度达到1.86 g/L。通过分子修饰获得Hpa-GalT的A186P/E146Q变体，使LNnT滴度提高到2.15 g/L。随后，lacZ、ugd、wcaJ、wecB等一系列与竞争通路相关的基因被删除，工程菌株的LNnT滴度为2.23 g/L。优化LNnT运输和利用后，工程菌株AH31摇瓶发酵的LNnT产量为2.93 g/L。在5-L的发酵罐中，最大滴度为总LNnT 32.6 g/L，胞外LNnT 27.1 g/L，产率为0.52 g/L/h。总之，本研究提供了一种新的β-1,4-半乳糖转移酶，为LNnT的高效生物合成奠定了基础。

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引用次数: 0

Whole-cell biosensors with regulatory circuits based on a synthetic dual-input promoter enabling the highly sensitive detection of ultra-trace Cd2+ 具有基于合成双输入启动子的调节电路的全细胞生物传感器，能够高度灵敏地检测超痕量Cd2+

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2026-03-01 Epub Date: 2025-09-11 DOI: 10.1016/j.synbio.2025.09.008

Yuke Qin , Shuting Hu , Yibin Yang , Xiaoqiang Jia

The research on whole-cell biosensors tailored for trace and ultra-trace detection remains limited and the biosensors based on natural bacterial heavy metal resistance mechanisms have common issues of low sensitivity. In this study, we designed a promoter P_T7-cadO using the Cadmium ions (Cd²⁺)-binding protein binding site cadO with the T7 promoter at first and constructed a single-input whole-cell biosensor, which was named CP100, whose detection limit for Cd²⁺ met the WHO requirement, yet its response and sensitivity were quite low. We further introduced the lacI and lac operator (lacO) as the signal amplifier to construct a dual-input promoter P_{T7-cadO-lacO-cadO} and developed a biosensor named LC100, whose response and sensitivity were significantly improved, but background leakage became a new problem. Then we redesigned the gene circuit based on the regulatory circuit LCPM-2, which has the structure of “CadR-P_J23100-P_{T7-cadO-lacO-cadO}-mRFP1-LacI”, with the LacI protein as the autoregulatory negative feedback model and finally obtained the biosensor named LC100-2, which achieved the detection of ultra-trace Cd²⁺ (0.00001–0.02 nM), with the sensitivity of 3748.22 times that of CP100. Moreover, LC100-2 demonstrated excellent specificity to Cd²⁺ among four other divalent metal ions and good anti-interference capability in the mixed divalent metal ions system. The results of the real water sample tests demonstrated that precise quantitative detection of Cd²⁺ with a final concentration of 0.001–0.02 nM could be achieved by adding only a small volume of the sample (1 μL). This finding showed promising application potential in the field of trace detection. Additionally, the regulatory circuit LCPM-2 based on the unique dual-input promoter enhanced responses and reduced background leakage simultaneously, thereby providing an innovative strategy for the design and simplification of biosensors’ circuits.

针对痕量和超痕量检测的全细胞生物传感器的研究仍然有限，基于天然细菌重金属抗性机制的生物传感器普遍存在灵敏度低的问题。本研究首先利用镉离子（Cd2+）结合蛋白结合位点cadO与T7启动子设计了启动子PT7-cadO，构建了单输入全细胞生物传感器CP100，该传感器对Cd2+的检出限符合WHO要求，但其响应和灵敏度较低。我们进一步引入lacI和lac算子（lacO）作为信号放大器，构建双输入启动子PT7-cadO-lacO-cadO，并开发了LC100生物传感器，其响应和灵敏度得到了显著提高，但背景泄漏成为新的问题。然后基于结构为“CadR-PJ23100-PT7-cadO-lacO-cadO-mRFP1-LacI”的调控电路LCPM-2重新设计基因电路，以LacI蛋白为自调控负反馈模型，最终获得命名为LC100-2的生物传感器，实现了超微量Cd2+ (0.00001-0.02 nM)的检测，灵敏度为CP100的3748.22倍。LC100-2对其他四种二价金属离子中的Cd2+具有良好的特异性，在混合二价金属离子体系中具有良好的抗干扰能力。实际水样测试结果表明，只需加入少量样品（1 μL），就可以实现对Cd2+的精确定量检测，最终浓度为0.001 ~ 0.02 nM。这一发现在痕量检测领域具有广阔的应用前景。此外，基于独特双输入启动子的调控电路LCPM-2增强了响应，同时减少了背景泄漏，从而为生物传感器电路的设计和简化提供了一种创新策略。

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引用次数: 0

Rapid detection of genotoxicity employing an engineered high-sensitivity and tight SOS-response biosensor in Escherichia coli MG1655 利用工程化的高灵敏度、紧密sos反应生物传感器快速检测大肠杆菌MG1655的遗传毒性

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2026-03-01 Epub Date: 2025-06-27 DOI: 10.1016/j.synbio.2025.06.011

Xin Yuan , Wenliang Hao , Chong Zhang , Yizhi Ji , Yi Wang

Industrial wastewater and pesticide residues contain numerous genotoxic compounds that pose serious health risks by causing DNA damage and potentially triggering carcinogenesis. Conventional analytical methods such as chromatography and mass spectrometry are limited by high costs, technical complexity, and lengthy procedures, making them unsuitable for rapid, large-scale screening. To address this need, we developed a fluorescence-based SOS-responsive whole-cell biosensor (SRWCB) by integrating the P_sulA promoter with superfolder GFP (sfGFP). Through directed evolution of the lexA binding motif in P_sulA, we obtained an optimized variant (P55) with significantly enhanced sensitivity—being 8.5 times more sensitive than the commonly used P_umuDC system—while maintaining rapid response kinetics. Further engineering via antisense transcription improved biosensor tightness without compromising induction efficiency. The final construct (SRWCB-55-J23119) successfully detected genotoxicants in industrial wastewater, demonstrating its practical applicability. This study presents a sensitive, rapid, and robust biosensor platform for high-throughput environmental genotoxicity monitoring.

工业废水和农药残留含有大量的基因毒性化合物，这些化合物会造成DNA损伤并可能引发致癌，从而对健康构成严重威胁。传统的分析方法，如色谱法和质谱法，受成本高、技术复杂和程序长等限制，不适合快速、大规模的筛选。为了满足这一需求，我们通过将PsulA启动子与超级文件夹GFP （sfGFP）整合，开发了基于荧光的sos响应全细胞生物传感器（SRWCB）。通过对PsulA中lexA结合基序的定向进化，我们获得了一个优化的变体（P55），其灵敏度显著提高，是常用PumuDC系统的8.5倍，同时保持了快速的反应动力学。进一步工程通过反义转录提高了生物传感器的紧密性，而不影响诱导效率。最终构建的SRWCB-55-J23119成功检测了工业废水中的基因毒物，证明了其实用性。本研究提出了一种灵敏、快速、稳健的生物传感器平台，用于高通量环境遗传毒性监测。

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引用次数: 0

Systems metabolic engineering of dual pathways with stage-specific activation for high-level 5-aminolevulinic acid production in Escherichia coli 大肠杆菌生产高水平5-氨基乙酰丙酸的双途径与阶段特异性激活的系统代谢工程

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2026-03-01 Epub Date: 2025-09-08 DOI: 10.1016/j.synbio.2025.09.007

Yanghao Liu , Changgeng Li , Yu Miao , Zhichao Chen , Tang'en Shi , Ling Ma , Siyu Zhao , Zichen Yu , Lanxiao Li , Qingyang Xu

5-Aminolevulinic acid (5-ALA) is an important non-proteinogenic amino acid with wide applications in agriculture and medicine. To achieve high-level microbial production, this study established a dual-pathway reconstruction strategy in Escherichia coli by integrating the endogenous C5 pathway with an inducible exogenous C4 pathway. Multi-copy overexpression of gltX, hemA, and hemL, combined with enhanced glutamate supply and the introduction of a non-oxidative glycolysis pathway, effectively increased the C5 pathway flux and carbon efficiency, while product toxicity was significantly alleviated by strengthening efflux mechanisms and oxidative stress tolerance. A quorum sensing–based regulatory system was employed to dynamically regulate hemB expression, thereby balancing cell growth and 5-ALA biosynthesis, and a controlled glycine feeding strategy specifically activated the C4 pathway during the later fermentation stage, forming a stage-specific dynamic activation mechanism. Promoter regulation of sucC/sucD expression and enhanced endogenous PLP biosynthesis further stabilized the C4 flux. Fed-batch fermentation in a 5 L bioreactor resulted in a final 5-ALA titer of 37.34 g/L, demonstrating the industrial potential of this systems metabolic engineering strategy combining dual pathways and stage-specific activation control.

5-氨基乙酰丙酸（5-ALA）是一种重要的非蛋白质原性氨基酸，在农业和医学上有着广泛的应用。为了实现高水平的微生物生产，本研究通过整合内源性C5途径和可诱导的外源性C4途径，在大肠杆菌中建立了双途径重建策略。gltX、hemA和hemL的多拷贝过表达，结合谷氨酸供应的增加和非氧化糖酵解途径的引入，有效地增加了C5途径的通量和碳效率，同时通过加强外排机制和氧化应激耐受性，显著减轻了产物毒性。利用群体感应调控系统动态调控hemB表达，平衡细胞生长和5-ALA的生物合成，调控甘氨酸取食策略在发酵后期特异性激活C4通路，形成阶段性动态激活机制。启动子对sucC/sucD表达的调控和内源性PLP生物合成的增强进一步稳定了C4通量。在5l生物反应器中分批补料发酵，最终5- ala滴度为37.34 g/L，证明了该系统代谢工程策略结合双途径和阶段特异性激活控制的工业潜力。

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引用次数: 0

Integrated transcriptomic and metabolomic analyses uncover novel genetic targets for enhanced l-tryptophan production in Corynebacterium glutamicum 整合转录组学和代谢组学分析揭示了谷氨酸棒状杆菌增强l-色氨酸生产的新遗传靶点

IF 4.4 2区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Synthetic and Systems Biotechnology

Pub Date : 2026-03-01 Epub Date: 2025-09-29 DOI: 10.1016/j.synbio.2025.09.020

Yufei Dong , Rongsheng Gao , Nan Qin , Kunyu Liu , Youmeng Liu , Zhen Chen

Corynebacterium glutamicum is a promising microbial chassis for the industrial production of l-tryptophan, an essential amino acid with diverse applications and high market value. In our previous work, we constructed an l-tryptophan-overproducing C. glutamicum strain, TR26, through multiple rounds of rational metabolic engineering. Here, comparative transcriptomic and metabolomic analyses were conducted between TR26 and its progenitor MB001 to elucidate the underlying molecular mechanisms and identify potential bottlenecks for l-tryptophan production in TR26. Based on the differentially expressed genes identified, systematic gene overexpression and knockdown experiments led to the identification of two novel genetic targets, glnK and sugR. Specifically, the repression of glnK and overexpression of sugR in TR26 increased the l-tryptophan titer by 10.3 % and 16.5 % in fed-batch fermentation, and the yield by 7.2 % and 20.2 %, respectively. Further transcription profiling and intracellular metabolite analysis indicated that these improvements were associated with altered nitrogen metabolism, more efficient allocation of cellular resources, and enhanced supply of phosphoenolpyruvate (PEP), a key precursor in aromatic amino acid biosynthesis. This study expands our understanding of the regulation mechanisms governing l-tryptophan synthesis in C. glutamicum and provides valuable insights for further optimization of industrial cell factories.

l-色氨酸是一种用途广泛、具有较高市场价值的必需氨基酸，谷氨酸棒状杆菌是工业生产l-色氨酸的理想微生物基质。在之前的工作中，我们通过多轮合理的代谢工程，构建了过量生产l-色氨酸的谷氨酸C.菌株TR26。本研究对TR26及其祖基因MB001进行了转录组学和代谢组学比较分析，以阐明TR26产生l-色氨酸的潜在分子机制，并确定TR26产生l-色氨酸的潜在瓶颈。在鉴定出差异表达基因的基础上，通过系统的基因过表达和敲低实验，鉴定出两个新的基因靶点glnK和sugR。具体来说，抑制glnK和TR26过表达糖r可使l-色氨酸滴度提高10.3%和16.5%，产量分别提高7.2%和20.2%。进一步的转录分析和细胞内代谢物分析表明，这些改善与氮代谢的改变、细胞资源的更有效分配和磷酸烯醇丙酮酸（PEP）的供应增加有关，PEP是芳香氨基酸生物合成的关键前体。本研究扩展了我们对谷氨酸酵母l-色氨酸合成调控机制的理解，并为进一步优化工业细胞工厂提供了有价值的见解。

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引用次数: 0