Metabolic engineering最新文献_第7页

Multi-layered transcriptional and post-translational fine-tuning of metabolic pathways for overproduction of L-valine l -缬氨酸过量产生的代谢途径的多层转录和翻译后微调。

IF 6.8 1区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Metabolic engineering

Pub Date : 2026-01-01 Epub Date: 2025-09-18 DOI: 10.1016/j.ymben.2025.09.005

Xutao Lang , Wenwen Yu , Xuewen Zhu , Xianhao Xu , Yanfeng Liu , Jianghua Li , Guocheng Du , Jian Chen , Xueqin Lv , Long Liu

L-valine is an essential amino acid widely used in the food, pharmaceutical, and animal feed industries. Currently, engineering microbial cell factories to produce L-valine from low-cost feedstocks has emerged as a leading strategy. However, there is still a lack of an L-valine-producing strain that simultaneously exhibits high titer, high yield, and high productivity. The metabolic engineering strategies reported for L-valine biosynthesis primarily rely on conventional unidimensional, transcriptional-level modifications, which limit fine-tuning and do not provide comprehensive, multi-layered regulation. In this study, we constructed an Escherichia coli hyperproducer of L-valine by multiplexed transcriptional and post-translational fine-tuning of metabolic pathways. Initially, the transcriptional repression in L-valine synthetic pathway was eliminated by using promoter engineering strategy. Then, the “push-pull-inhibit” and transport engineering strategies were used to improve L-valine accumulation, achieving a flask fermentation titer of 21.6 g/L and a productivity of 0.45 g/L/h. Subsequently, we rationally designed membraneless organelles (MLOs) to enable spatial regulation of metabolic biosynthesis, which enhanced the targeted recruitment of dihydroxy-acid dehydratase and branched-chain amino acid aminotransferase. This spatial reorganization led to a 95.6 % increase in productivity, reaching 0.88 g/L/h. Finally, the best-performing strain produced 90.6 g/L L-valine in a 3-L bioreactor at 28 h, with a yield of 0.48 g/g glucose and a productivity of 3.24 g/L/h. To the best of our knowledge, this represents the highest L-valine productivity achieved to date. Our strategy provides a practical and effective approach for advancing microbial amino acid biosynthesis by multi-layered transcriptional and post-translational regulation of metabolic pathways.

l -缬氨酸是一种必需氨基酸，广泛应用于食品、制药和动物饲料行业。目前，工程微生物细胞工厂以低成本原料生产l -缬氨酸已成为一种主要策略。然而，仍然缺乏一种l -缬氨酸生产菌株，同时表现出高滴度，高产率和高生产力。据报道，l-缬氨酸生物合成的代谢工程策略主要依赖于传统的一维转录水平修饰，这限制了微调，不能提供全面的、多层次的调控。在这项研究中，我们通过多重转录和翻译后的代谢途径微调构建了大肠杆菌l -缬氨酸高产菌。最初，利用启动子工程策略消除了l -缬氨酸合成途径中的转录抑制。然后，采用“推-拉-抑制”和运输工程策略提高L-valine积累，达到了21.6 g/L的烧瓶发酵滴度和0.45 g/L/h的产率。随后，我们合理设计无膜细胞器（MLOs），实现代谢生物合成的空间调控，增强了二羟基酸脱水酶和支链氨基酸转氨酶的靶向募集。这种空间重组使生产力提高了95.6%，达到0.88 g/L/h。结果表明，最佳菌株在3-L生物反应器中28h产L-valine 90.6 g/L，产糖量为0.48 g/g，产率为3.24 g/L/h。据我们所知，这代表了迄今为止最高的l -缬氨酸生产力。我们的策略提供了一种实用有效的方法，通过多层转录和翻译后代谢途径的调节来推进微生物氨基酸的生物合成。

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引用次数: 0

Metabolic and enzyme rewiring enables high-production of vanillin in unconventional yeast 代谢和酶重组使非常规酵母的香兰素高产成为可能。

IF 6.8 1区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Metabolic engineering

Pub Date : 2026-01-01 Epub Date: 2025-10-09 DOI: 10.1016/j.ymben.2025.10.002

Yan Guo , Liyang Zhou , Wanshu Lai , Zhilan Qian , Haishuang Yu , Menghao Cai

Vanillin is an aromatic flavor compound widely used in the food, pharmaceutical, and cosmetic industries. Microbial biosynthesis offers a sustainable alternative to traditional plant extraction and chemical synthesis; however, the susceptibility of vanillin to redox reactions and the weak enzyme activity in cells severely limit the vanillin production capacity by microbial biosynthesis. This study presents the first successful attempt at de novo synthesis of vanillin in the unconventional yeast Komagataella phaffii. The initial titer was quite low (0.5 mg/L), but removal of 14 endogenous oxidoreductases to block vanillin conversion resulted in an 11.1-fold improvement in vanillin production. The combination of pathway rewiring and cofactor (nicotinamide adenine dinucleotide phosphate [NADPH] and S-adenosylmethionine) regeneration redirected the metabolic flux toward vanillin synthesis and achieved a further 19.9-fold improvement in vanillin production. Rational rewiring of the rate-limiting enzyme, caffeic acid O-methyltransferase (NtCOMT), generated a dominant mutant NtCOMT^N312A/H315N from 70 variants, which promoted activity by 49.7 % and prevented intermediate accumulation. These strategies eventually enabled the co-coupling of de novo biosynthesis and caffeic acid conversion, achieving the highest reported production of vanillin (1055.9 mg/L) by K. phaffii fermentation in a bioreactor. These findings highlight the potential of unconventional yeast as a chassis host for aromatic aldehyde synthesis and the construction of a versatile microbial platform for the production of carbonyl compounds.

香兰素是一种芳香香料化合物，广泛应用于食品、制药和化妆品行业。微生物生物合成为传统的植物提取和化学合成提供了可持续的替代方案；然而，由于香兰素对氧化还原反应的敏感性和细胞内酶活性较弱，严重限制了微生物合成香兰素的生产能力。本研究首次成功尝试在非常规酵母法菲酵母中重新合成香兰素。初始滴度很低（0.5 mg/L），但去除14个内源性氧化还原酶以阻断香兰素转化，使香兰素产量提高了11.1倍。途径重组和辅助因子（烟酰胺腺嘌呤二核苷酸磷酸[NADPH]和s -腺苷蛋氨酸）再生的结合将代谢通量转向香兰素合成，并使香兰素产量进一步提高19.9倍。通过对限速酶咖啡酸o -甲基转移酶（NtCOMT）的合理重新连接，从70个突变体中产生了显性突变体NtCOMTN312A/H315N，其活性提高了49.7%，并阻止了中间积累。这些策略最终实现了新生物合成和咖啡酸转化的共偶联，在生物反应器中通过K. phaffii发酵实现了最高的香兰素产量（1055.9 mg/L）。这些发现突出了非传统酵母作为芳香醛合成的基础宿主和构建生产羰基化合物的多功能微生物平台的潜力。

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引用次数: 0

Metabolic engineering of Acinetobacter baylyi ADP1 for efficient utilization of pentose sugars and production of glutamic acid 贝氏不动杆菌ADP1代谢工程对戊糖的高效利用和谷氨酸的生产。

IF 6.8 1区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Metabolic engineering

Pub Date : 2026-01-01 Epub Date: 2025-10-09 DOI: 10.1016/j.ymben.2025.10.001

Jin Luo, Elena Efimova, Ville Santala, Suvi Santala

Efficient utilization of pentose sugars is critical for advancing sustainable biomanufacturing using lignocellulose. However, many host strains capable of consuming glucose and lignin-derived monomers are unable to utilize pentose sugars. Here, we engineered Acinetobacter baylyi ADP1 for the utilization of D-xylose and L-arabinose. We first modelled different pentose utilization pathways using flux balance analysis to choose the most optimal pathway. A marker-free approach combining transformation and selection facilitated the integration of the pentose catabolic gene clusters of the selected Weimberg pathway into the A. baylyi genome, generating strains capable of efficiently utilizing both D-xylose and L-arabinose as sole carbon sources without any additional engineering or adaptation. For D-xylose, the cells achieved the highest growth rate (μ = 0.73 h⁻¹) reported to date for non-native hosts engineered for pentose utilization. For L-arabinose, a growth rate of μ = 0.40 h⁻¹ was achieved, which also surpassed the growth rate on a native substrate of A. baylyi, glucose (μ = 0.37 h⁻¹). Importantly, pentose utilization occurred simultaneously with glucose utilization. We then applied metabolic flux analysis using ¹³C labeled xylose to reveal D-xylose metabolism in the engineered strain. To demonstrate the potential for bioproduction, L-glutamate was selected as a target compound. Deletion of sucAB and gabT, and the overexpression of gdhA enabled L-glutamate production. With the engineered strain, a carbon yield of 34 % during co-utilization with succinate and 70 % via whole-cell catalysis using resting cells was achieved. Notably, L-glutamate production directly from industrially relevant hemicellulose hydrolysate was demonstrated. This study establishes a robust platform for pentose utilization and bioproduction in A. baylyi ADP1 and highlights the potential for metabolic optimization.

戊糖的有效利用对于推进木质纤维素的可持续生物制造至关重要。然而，许多能够消耗葡萄糖和木质素衍生单体的宿主菌株不能利用戊糖。在这里，我们设计了利用d -木糖和l -阿拉伯糖的baylyi不动杆菌ADP1。首先利用通量平衡分析对不同戊糖利用途径进行建模，选择最优途径。结合转化和选择的无标记方法促进了Weimberg途径的戊糖分解代谢基因簇整合到A. baylyi基因组中，产生了能够有效利用d -木糖和l -阿拉伯糖作为唯一碳源的菌株，而无需任何额外的工程或适应。对于d -木糖，细胞的生长速度（μ=0.73 h-1）是迄今为止报道的用于戊糖利用的非原生宿主中最高的。l -阿拉伯糖的生长速度为μ=0.40 h-1，也超过了在天然底物葡萄糖上的生长速度（μ=0.37 h-1）。重要的是，戊糖利用与葡萄糖利用同时发生。然后用13C标记木糖进行代谢通量分析，揭示工程菌株d -木糖代谢。为了证明生物生产的潜力，选择l -谷氨酸作为目标化合物。缺失sucAB和gabT以及过表达gdhA使l -谷氨酸产生。在与琥珀酸盐共利用的过程中，该工程菌株的碳产量为34%，在静息细胞的全细胞催化下，碳产量为70%。值得注意的是，从工业相关的半纤维素水解物中直接生产l -谷氨酸得到了证明。本研究为baylyi ADP1的戊糖利用和生物生产建立了一个强大的平台，并强调了代谢优化的潜力。

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引用次数: 0

From soil to biomanufacturing: Systems-driven metabolic pathway rewiring in non-model bacteria for gram-scale antibiotic production 从土壤到生物制造：系统驱动的代谢途径在非模式细菌中重新布线，用于克量级的抗生素生产

IF 6.8 1区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Metabolic engineering

Pub Date : 2026-01-01 Epub Date: 2025-11-04 DOI: 10.1016/j.ymben.2025.11.001

Tingfeng Cheng , Suihao Yan , Min Xu , Lei Zhao

Microbial natural products (NPs) are a pivotal reservoir for drugs used in human health and agriculture. Andrimid, a polyketide-non-ribosomal peptide hybrid antibiotic inhibiting bacterial acetyl-CoA carboxylase, shows enormous potential in antibiotic drug development to mitigate antimicrobial resistance. However, industrial-scale manufacturing and downstream development of andrimid are largely prohibited due to its milligram level production in microorganisms. Herein, using an integrative multi-omics approach, we improved the yield of andrimid remarkably from milligram to gram level in a non-model environmental soil bacterium, Erwinia persicina BST187, isolated from the rhizosphere of tomato. Systematic reprogramming of the pathways for carbon source uptake, competing metabolites biosynthesis, supply of essential building blocks including phenylalanine, glycine, valine and malonyl-CoA and cofactor biosynthesis using CRIPSR/Cas9 based gene editing tools, coupled with fine-tuning the transcription of the biosynthetic genes of andrimid, resulted in the generation of the optimal producer, G17. Combined with fermentation optimization, andrimid was produced to a highest level of 1099.42 mg/L with a productivity of 15.3 mg/L/h using a 5 L bioreactor, representing a 628-fold increase compared to the parental strain. This study showcases the genome wide engineering of non-model bacteria and generates a plasmid- and inducer-free E. persicina strain for high-level andrimid production, providing a blueprint for systems-driven metabolic engineering of complex bioactive NPs for biomanufacturing.

微生物天然产物（NPs）是用于人类健康和农业的药物的关键储存库。Andrimid是一种抑制细菌乙酰辅酶a羧化酶的聚酮-非核糖体多肽混合抗生素，在抗生素药物开发中显示出巨大的潜力。然而，工业规模的制造和下游开发在很大程度上是被禁止的，因为它的毫克水平的微生物生产。本研究利用综合多组学方法，从番茄根际分离的非模式环境土壤细菌Erwinia persicina BST187中显著提高了雌雄同体的产量，从毫克到克水平。利用基于CRIPSR/Cas9的基因编辑工具，系统地对碳源吸收、竞争性代谢物生物合成、苯丙氨酸、甘氨酸、缬氨酸和丙二酰辅酶a等必需构建块的供应以及辅助因子的生物合成途径进行了重编程，再加上对雄酰胺生物合成基因的转录进行了微调，最终产生了最佳生产者G17。结合发酵优化，在5 L的生物反应器中，雄酰胺的最高产率为1099.42 mg/L，产率为15.3 mg/L/h，比亲本菌株提高了628倍。本研究展示了非模式细菌的全基因组工程，并产生了一种无质粒和无诱导性的persicina菌株，用于高水平的雄酰胺生产，为生物制造中复杂生物活性NPs的系统驱动代谢工程提供了蓝图。

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引用次数: 0

Systematic rewiring of Bacillus subtilis for efficient de novo biosynthesis of the neuroprotectant cytidine-5′-diphosphocholine 枯草芽孢杆菌系统重新布线，有效地重新合成神经保护剂胞苷-5'-二磷酸胆碱。

IF 6.8 1区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Metabolic engineering

Pub Date : 2026-01-01 Epub Date: 2025-09-11 DOI: 10.1016/j.ymben.2025.09.004

Shaomei Yang , Xu Feng , Hao Wei , Yanshuang Wang , Shouying Fu , Xiuzhen Gao , Qinyuan Ma

Cytidine-5′-diphosphocholine (CDP-choline) is a crucial neuroprotective agent. Current industrial production relies on chemical and enzymatic methods that face inherent sustainability challenges and share a dependence on the costly precursor, cytidine monophosphate (CMP). Here, we report the systems metabolic engineering of Bacillus subtilis for the efficient, de novo biosynthesis of CDP-choline from glucose, completely obviating the need for CMP. A synthetic pathway was first established by introducing heterologous choline kinase and phosphocholine cytidylyltransferase. Subsequently, a multi-module engineering strategy was implemented, focusing on enhancing precursor supply and redirecting carbon metabolism. This involved systematically optimizing choline uptake by overexpressing the transporter OpuD and deleting the transcriptional repressor opcR, fortifying the cytidine triphosphate (CTP) pool by overexpressing feedback-resistant CTP synthase gene pyrG^E156K, and deleting the transcriptional repressor pyrR along with other pyrimidine nucleotide consumption genes, and channeling carbon flux towards the TCA cycle by reducing pyruvate and malate consumption. The final engineered strain achieved a titer of 4.79 ± 0.24 g/L CDP-choline in a 5 L fed-batch bioreactor, with a high specific yield of 149.0 ± 5.8 mg/g DCW. Notably, the process exhibited a highly advantageous intracellular accumulation of 92.7 %, which simplifies downstream purification. This study represents the first successful demonstration of CDP-choline production from simple sugars in a microbial host, establishing a robust and economically competitive platform for its industrial manufacture.

胞苷-5′-二磷酸胆碱（cdp -胆碱）是一种重要的神经保护剂。目前的工业生产依赖于化学和酶的方法，这些方法面临着固有的可持续性挑战，并且依赖于昂贵的前体，单磷酸胞苷（CMP）。在这里，我们报道了枯草芽孢杆菌的系统代谢工程，以有效地从葡萄糖中重新合成cdp -胆碱，完全消除了对CMP的需要。通过引入外源胆碱激酶和磷酸胆碱胞基转移酶，初步建立了一条合成途径。随后，实施了多模块工程策略，重点是增加前体供应和重定向碳代谢。这包括通过过表达转运体OpuD和删除转录抑制因子opcR来系统地优化胆碱摄取，通过过表达反馈抗性CTP合成酶基因pyrGE156K来强化三磷酸胞苷（CTP）库，并删除转录抑制因子pyrR和其他嘧啶核苷酸消耗基因，并通过减少丙酮酸和苹果酸消耗来引导碳通量进入TCA循环。在5 L补料间歇式生物反应器中，最终工程菌株的滴度为4.79±0.24 g/L，比产率为149.0±5.8 mg/g DCW。值得注意的是，该工艺具有92.7%的胞内富集率，简化了下游纯化。这项研究首次成功地展示了在微生物宿主中由单糖生产cdp -胆碱，为其工业生产建立了一个强大的、具有经济竞争力的平台。

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引用次数: 0

Reconstruction of a resource balance analysis model of Clostridium thermocellum examines the metabolic cost of glycolytic and cellulosome enzymes 热梭菌资源平衡分析模型的重建研究了糖酵解和纤维素酶的代谢成本。

IF 6.8 1区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Metabolic engineering

Pub Date : 2026-01-01 Epub Date: 2025-09-08 DOI: 10.1016/j.ymben.2025.09.001

Thomas C. Willis , Wheaton L. Schroeder , Daven B. Khana , Xuejun Qian , Sanjeev Dahal , Daniel Amador-Noguez , Costas D. Maranas

Clostridium thermocellum is an increasingly well-studied organism with considerable advantages for consolidated bioprocessing towards ethanol production. Here, a genome-scale resource balance analysis (RBA) model of C. thermocellum, ctRBA, is reconstructed based on a recently published stoichiometric model (iCTH669), global proteomics, and ¹³C MFA datasets to analyze proteome allocation and the burden imposed on metabolism with regard to ethanol yield and titer. Glycolytic and fermentation enzyme concentrations were accurately quantified by the model, with glyceraldehyde-3-phosphate dehydrogenase (GAPDH), phosphoglycerate kinase (PGK), and acetaldehyde-alcohol dehydrogenase (AdhE) having predicted and measured higher concentrations relative to other enzymes in glycolysis and fermentation. The metabolic burden associated with the formation of the cellulosome, the enzyme complex responsible for carbon source degradation and solubilization, was assessed and found to be consequential in constraining ethanol yield and titer, but not biomass formation. Putative enzyme substitution strains were modeled, with each strain replacing a single enzyme in C. thermocellum with a variant that uses more favorable cofactors. Strains substituting GAPDH and phosphofructokinase (PFK) predicted 30 % and 86 % increases in maximum theoretical ethanol yield and titer, respectively, a result unavailable to typical stoichiometric modeling. Model ctRBA acts as a predictive tool for assessing the effect of genetic perturbations on proteome allocation and ethanol yield and titer.

热胞梭菌是一种研究越来越深入的生物，在乙醇生产的强化生物处理中具有相当大的优势。本文基于最近发表的化学计量模型（iCTH669）、全球蛋白质组学和13C MFA数据集，重建了热cellum （ctRBA）的基因组尺度资源平衡分析（RBA）模型，以分析蛋白质组分配以及乙醇产量和滴度对代谢的影响。通过该模型，糖酵解和发酵酶的浓度被精确量化，甘油醛-3-磷酸脱氢酶（GAPDH）、磷酸甘油激酶（PGK）和乙醛-醇脱氢酶（AdhE）预测和测量的浓度相对于糖酵解和发酵中的其他酶更高。与纤维素（负责碳源降解和溶解的酶复合体）的形成相关的代谢负担被评估，并发现在限制乙醇产量和滴度方面是重要的，但不是生物量的形成。对假定的酶替代菌株进行建模，每个菌株用使用更有利的辅因子的变体替换C. thermocellum中的单个酶。取代GAPDH和磷酸果糖激酶（PFK）的菌株预测最大理论乙醇产量和滴度分别提高30%和86%，这一结果无法通过典型的化学计量模型得到。ctRBA模型作为一种预测工具，用于评估遗传扰动对蛋白质组分配、乙醇产量和滴度的影响。

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引用次数: 0

Lipid accumulation in nitrogen and phosphorus-limited yeast is caused by less growth-related dilution 氮磷限制酵母中的脂质积累是由较少的生长相关稀释引起的

IF 6.8 1区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Metabolic engineering

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

Xi Li , Daniel R. Weilandt , Felix C. Keber , Arjuna M. Subramanian , Shayne R. Loynes , Christopher V. Rao , Yihui Shen , Martin Wühr , Joshua D. Rabinowitz

Oleaginous yeasts are used commercially to produce oleochemicals and hold potential also for biodiesel production. In response to nitrogen or phosphorous limitation, oleaginous yeasts accumulate lipids in the form of triacylglycerols. Previous work has investigated potential mechanisms by which nutrient limitation induces lipid biosynthesis without verifying whether lipid biosynthesis flux is actually enhanced. Here we show, using ¹³C-glucose tracing, that in nitrogen or phosphorous limitation, lipid accumulation occurs without consistent increases in biosynthetic flux. Instead, the main driver of increased lipid pools is decreased growth-related dilution. This conclusion holds across two divergent oleaginous yeasts: Rhodotorula toruloides and Yarrowia lipolytica. Quantitative proteomics shows a substantial proteome reallocation in response to nitrogen and phosphorous limitation, with ribosomal proteins strongly downregulated, while lipid enzymes are preserved but not consistently upregulated in absolute quantity. Thus, nutrient limitation, rather than triggering greatly enhanced lipid synthesis, results in roughly sustained lipid enzyme levels and biosynthetic flux. Due to slower lipid dilution by cell division, this suffices to drive marked lipid accumulation.

产油酵母在商业上用于生产油脂化学品，也具有生产生物柴油的潜力。由于氮或磷的限制，产油酵母以三酰基甘油的形式积累脂质。以前的工作研究了营养限制诱导脂质生物合成的潜在机制，但没有验证脂质生物合成通量是否实际上增强。在这里，我们表明，使用13c -葡萄糖示踪，在氮或磷限制下，脂质积累发生在生物合成通量不一致增加的情况下。相反，脂质池增加的主要驱动因素是与生长相关的稀释度降低。这一结论适用于两种不同的产油酵母：环形红酵母和多脂耶氏酵母。定量蛋白质组学显示，在氮和磷的限制下，蛋白质组发生了实质性的再分配，核糖体蛋白被强烈下调，而脂质酶被保留，但在绝对数量上不一致上调。因此，营养限制，而不是触发大大增强脂质合成，导致大致维持脂质酶水平和生物合成通量。由于细胞分裂对脂质稀释较慢，这足以驱动显著的脂质积累。

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引用次数: 0

Developing a Redox Imbalance Forces Drive (RIFD) strategy and its application in L-threonine production 氧化还原不平衡力驱动（RIFD）策略及其在l -苏氨酸生产中的应用。

IF 6.8 1区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Metabolic engineering

Pub Date : 2025-11-01 Epub Date: 2025-08-05 DOI: 10.1016/j.ymben.2025.07.014

Xin Jin , Ruxin Hao , Hannuo Shen , Zhu Liu , Sumeng Wang , Qingsheng Qi , Quanfeng Liang

The design of synthetic driving forces for biosynthetic pathway is crucial for directing carbon flux toward the target product. Optimizing cellular redox status is one of the key strategies for constructing microbial cell factories. In this study, we attempt to create a novel redox imbalance force-driven (RIFD) strategy to direct carbon flow toward the target synthetic pathway. Initially, we increased the NADPH pool through a strategy of “open source and reduce expenditure” employing four approaches to achieve excessive NADPH levels and growth inhibition: (I) the expression of cofactor-converting enzymes, (II) the expression of heterologous cofactor-dependent enzymes, (III) the expression of enzymes involved in the NADPH synthesis pathway, and (IV) reduced NADPH wastage by knocking down non-essential genes that consume NADPH in vivo. Next, multiple automated genome engineering (MAGE) techniques were employed to evolve redox-imbalanced engineered strains and drive the metabolic flux to L-threonine production. Finally, we developed a NADPH and L-threonine dual-sensing biosensor, combined it with Fluorescence-Activated Cell Sorting (FACS), and a high-yield (0.65 g/g) L-threonine-producing strain with a titer of 117.65 g L⁻¹ was obtained. This research presents a general approach to increasing the production of cofactor-related products. Utilizing redox imbalance forces to drive metabolic flow toward the target product, it is possible to increase production while simultaneously restoring cell growth.

生物合成途径合成驱动力的设计是将碳通量导向目标产物的关键。优化细胞氧化还原状态是构建微生物细胞工厂的关键策略之一。在本研究中，我们试图创建一种新的氧化还原不平衡力驱动（RIFD）策略，将碳流导向目标合成途径。最初，我们通过“开源和减少支出”的策略增加了NADPH池，采用四种方法来实现过量的NADPH水平和生长抑制：(I)表达辅酶转换酶，（II）表达异源辅酶依赖酶，（III）表达参与NADPH合成途径的酶，（IV）通过敲除体内消耗NADPH的非必需基因来减少NADPH的浪费。接下来，采用多种自动化基因组工程（MAGE）技术来进化氧化还原不平衡的工程菌株，并驱动l -苏氨酸生产的代谢通量。最后，我们开发了一种NADPH和L-苏氨酸双传感生物传感器，并将其与荧光活化细胞分选（FACS）技术结合，获得了一株L-苏氨酸高产菌株（0.65 g/g），滴度为117.65 g/L。本研究提出了增加辅因子相关产品生产的一般方法。利用氧化还原不平衡力来驱动代谢流向目标产物，有可能在增加产量的同时恢复细胞生长。

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引用次数: 0

Quantifying supply and demand in the pea aphid-Buchnera symbiosis reveals the metabolic Achilles’ heels of this interaction 定量的供应和需求在豌豆蚜虫- buchnera共生揭示了代谢的阿基里斯之踵，这种相互作用。

IF 6.8 1区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Metabolic engineering

Pub Date : 2025-11-01 Epub Date: 2025-07-26 DOI: 10.1016/j.ymben.2025.07.011

Léo Gerlin, Karen Gaget, Garance Lapetoule, Yohann Quivet, Patrice Baa-Puyoulet, Isabelle Rahioui, Mélanie Ribeiro Lopes, Pedro Da Silva, Federica Calevro, Hubert Charles

Many herbivorous insects feed on unbalanced diets and rely on bacterial endosymbionts to meet all their nutritional needs. This is the case for the pea aphid (Acyrthosiphon pisum), a plant pest whose remarkable growth and reproductive capacities cannot be sustained by its sole nutritional resource, the plant phloem sap, and which relies on a symbiotic relationship maintained over millions of years with the intracellular bacterium Buchnera aphidicola for the biosynthesis of amino acids and vitamins. Exploiting original experimental data and metabolic reconstructions, we have built a quantitative genome-scale metabolic model of B. aphidicola and used it to quantify amino acid exchanges between the bacterium and its host. We found metabolites that can rewire pathways, influencing the balance between selfish (growth-focused) and mutualist (amino acid synthesis) behavior. Among the products synthesized by Buchnera, phenylalanine, tyrosine and leucine are the main matter sinks and consume more than 60 % of imported glucose and serine. Finally, we compared the predicted bacterial supply to the aphid demand in amino acids. We found that the pea aphid may efficiently regulate its symbiont population density depending on its metabolic requirements, but that embryos are quantitatively not self-sustaining, with embryonic bacteria supply falling short of demand by 50 %. Overall, our study highlights candidate compounds and pathways to target for destabilizing this symbiosis or predicting its resilience to environmental or nutritional perturbations.

许多草食性昆虫以不平衡的饮食为食，依靠细菌内共生体来满足它们所有的营养需求。豌豆蚜虫（Acyrthosiphon pisum）就是这种情况，这种植物害虫的显著生长和繁殖能力不能靠其唯一的营养来源——植物韧皮部汁液来维持，它依赖于与细胞内细菌蚜虫（Buchnera aphidicola）维持了数百万年的共生关系来合成氨基酸和维生素。利用原始实验数据和代谢重建，我们建立了一个定量的蚜虫基因组尺度的代谢模型，并利用它来量化细菌与宿主之间的氨基酸交换。我们发现代谢物可以重新连接通路，影响自私（以生长为中心）和互惠（氨基酸合成）行为之间的平衡。在Buchnera合成的产品中，苯丙氨酸、酪氨酸和亮氨酸是主要的物质汇，消耗了60%以上的进口葡萄糖和丝氨酸。最后，我们比较了预测的细菌供应和蚜虫对氨基酸的需求。我们发现豌豆蚜虫可以根据其代谢需求有效地调节其共生体的种群密度，但胚胎在数量上不能自我维持，胚胎细菌供应不足需求的50%。总的来说，我们的研究突出了候选化合物和途径，以破坏这种共生关系或预测其对环境或营养扰动的恢复能力。

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引用次数: 0

Metabolic engineering of Micromonospora for exploring useful natural products and phytobiotic interaction 利用小单孢子菌代谢工程探索有用的天然产物和植物共生相互作用

IF 6.8 1区生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Metabolic engineering

Pub Date : 2025-11-01 Epub Date: 2025-07-20 DOI: 10.1016/j.ymben.2025.07.005

Boncheol Gu, Jimin Lee, Duck Gyun Kim, Yu-jin Cha, Min-Kyu Oh

Micromonospora, a genus within the Actinobacteria phylum, is recognized for its prolific production of bioactive secondary metabolites. It has important applications in the pharmaceutical, biotechnology, and agricultural fields. Micromonospora is renowned for generating antibiotics, anticancer agents, immunosuppressants, and plant growth-promoting compounds, making it a primary subject in natural product research. Advances in genome sequencing and mining technologies have revealed numerous biosynthetic gene clusters, many of which remain unexplored, underscoring their vast, untapped biosynthetic potential. This review presents an in-depth summary of the role of Micromonospora in the discovery of novel bioactive compounds and their biotechnological and industrial applications. Furthermore, we discuss the plant-microbe interactions of Micromonospora, consolidating current knowledge from its historical discovery to recent genomic insights, and outlines future research directions and challenges for optimizing the biotechnological potential of this promising yet underexploited microbial resource.

小单孢子菌是放线菌门中的一个属，以其多产的生物活性次生代谢物而闻名。它在制药、生物技术和农业领域有着重要的应用。小单孢子菌以产生抗生素、抗癌剂、免疫抑制剂和植物生长促进化合物而闻名，使其成为天然产物研究的主要课题。基因组测序和挖掘技术的进步揭示了许多生物合成基因簇，其中许多仍未被探索，强调了它们巨大的、未开发的生物合成潜力。本文综述了小单孢子菌在新型生物活性化合物的发现及其生物技术和工业应用中的作用。此外，我们讨论了小单孢菌的植物与微生物的相互作用，从其历史发现到最近的基因组见解，巩固了当前的知识，并概述了未来的研究方向和挑战，以优化这一有前途但尚未开发的微生物资源的生物技术潜力。

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引用次数: 0