Journal of biotechnology最新文献_第4页

Soft-sensing strategy for real-time biomass and poly(3-hydroxybutyrate) estimation in methanotrophic cultures using dissolved oxygen and base addition signals 利用溶解氧和碱添加信号实时估算甲烷营养培养物生物量和聚（3-羟基丁酸）的软测量策略。

IF 3.9 2区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Journal of biotechnology

Pub Date : 2025-12-16 DOI: 10.1016/j.jbiotec.2025.12.013

Seungyeon Cho , Kyutae Jung , Yoojin Shin , Inseo Kim , Seong-Hoon Jun , Hyo Jin Hong , Jeongwoo Lee , Jinwon Lee , Jeong-Geol Na

Real-time estimation of microbial biomass and product formation is crucial for effective monitoring and control of bioprocesses. In this study, we present a generalized soft-sensing algorithm that utilizes commonly available dissolved oxygen (DO) and base addition signals to estimate both total cell concentration and intracellular poly(3-hydroxybutyrate) (PHB) content. Total cell concentration was calculated using a DO-based oxygen balance equation, while PHB accumulation was inferred from a nitrogen balance framework, in which nitrogen consumption was estimated based on the frequency of base additions under pH-stat control. We validated the algorithm using methane-fed cultures of Methylocystis sp. MJC1, demonstrating high accuracy in predicting total biomass concentration across various media compositions and operational conditions, provided that ammonium was used as the nitrogen source. Notably, PHB concentrations were reliably estimated in real time by subtracting the non-PHB biomass (derived from base addition signals) from the DO-based total biomass. Although some deviations in PHB predictions were observed, these were attributed to imperfect correlations between nitrogen consumption and pH behavior. The method requires minimal data preprocessing and has low computational demands, making it well-suited for real-time applications in gas fermentation as well as other aerobic bioprocesses. This soft-sensing strategy offers a simple, robust, and cost-effective approach for online monitoring of microbial growth and product formation.

微生物生物量和产物形成的实时估计对于有效监测和控制生物过程至关重要。在这项研究中，我们提出了一种广义的软测量算法，该算法利用常用的溶解氧（DO）和碱添加信号来估计细胞总浓度和细胞内聚（3-羟基丁酸盐）（PHB）含量。总细胞浓度使用基于do的氧平衡方程计算，而PHB积累是从氮平衡框架推断的，其中氮消耗是根据ph值控制下的碱基添加频率估计的。我们用甲烷培养的Methylocystis sp. MJC1验证了该算法，证明了在不同培养基组成和操作条件下，只要使用铵作为氮源，该算法在预测总生物量浓度方面具有很高的准确性。值得注意的是，通过从基于do的总生物量中减去非PHB生物量（来自碱添加信号），可以可靠地实时估计PHB浓度。虽然在PHB预测中观察到一些偏差，但这些偏差归因于氮消耗与pH行为之间的不完全相关性。该方法需要最小的数据预处理和低计算需求，使其非常适合实时应用于气体发酵以及其他好氧生物过程。这种软测量策略为微生物生长和产品形成的在线监测提供了一种简单、稳健、经济的方法。

{"title":"Soft-sensing strategy for real-time biomass and poly(3-hydroxybutyrate) estimation in methanotrophic cultures using dissolved oxygen and base addition signals","authors":"Seungyeon Cho , Kyutae Jung , Yoojin Shin , Inseo Kim , Seong-Hoon Jun , Hyo Jin Hong , Jeongwoo Lee , Jinwon Lee , Jeong-Geol Na","doi":"10.1016/j.jbiotec.2025.12.013","DOIUrl":"10.1016/j.jbiotec.2025.12.013","url":null,"abstract":"<div><div>Real-time estimation of microbial biomass and product formation is crucial for effective monitoring and control of bioprocesses. In this study, we present a generalized soft-sensing algorithm that utilizes commonly available dissolved oxygen (DO) and base addition signals to estimate both total cell concentration and intracellular poly(3-hydroxybutyrate) (PHB) content. Total cell concentration was calculated using a DO-based oxygen balance equation, while PHB accumulation was inferred from a nitrogen balance framework, in which nitrogen consumption was estimated based on the frequency of base additions under pH-stat control. We validated the algorithm using methane-fed cultures of <em>Methylocystis</em> sp. MJC1, demonstrating high accuracy in predicting total biomass concentration across various media compositions and operational conditions, provided that ammonium was used as the nitrogen source. Notably, PHB concentrations were reliably estimated in real time by subtracting the non-PHB biomass (derived from base addition signals) from the DO-based total biomass. Although some deviations in PHB predictions were observed, these were attributed to imperfect correlations between nitrogen consumption and pH behavior. The method requires minimal data preprocessing and has low computational demands, making it well-suited for real-time applications in gas fermentation as well as other aerobic bioprocesses. This soft-sensing strategy offers a simple, robust, and cost-effective approach for online monitoring of microbial growth and product formation.</div></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"410 ","pages":"Pages 258-265"},"PeriodicalIF":3.9,"publicationDate":"2025-12-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145781438","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

AbrB and ResD negatively regulate locillomycin synthesis in Bacillus velezensis Bs916 by binding to the flanking sequences of transcription start sites abb和ResD通过与转录起始位点的侧翼序列结合，负向调控蓝芽孢杆菌Bs916中locillomycin的合成。

IF 3.9 2区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Journal of biotechnology

Pub Date : 2025-12-16 DOI: 10.1016/j.jbiotec.2025.12.010

Xiaohua Wang , HanWen Shen , Peng Zhang , Zhongzheng Ji , Xiaoxia Xu , Faxiang Wan , Xiulian Yin , Qingyun Bai , Liangcheng Du , Jiasong Jiang , Chuping Luo

Locillomycin, a cyclic lipopeptide produced by Bacillus velezensis Bs916, has been proven to have strong antimicrobial activity and low hemolytic activity. However, its low yield has become a significant obstacle for large-scale application. To identify the regulatory factors of locillomycin and obtain its high-yield strains through gene knockout, the transcriptional landscape of loc gene cluster was elucidated, and ResD and AbrB were identified as potential transcription regulators of the locillomycin biosynthesis gene cluster. EMSA and DNase Ⅰ footprinting results showed that AbrB tended to bind to A + T-rich fragments and exhibited extensive DNA-binding properties, being able to directly associate with the promoter, 5′ UTR, and coding region of the loc gene cluster. In contrast, the binding of ResD was confined to the promoter sequence upstream of the loc transcription start site, spanning from −480 to −456. To verify their impact on locillomycin production, mutants ΔresD, ΔabrB and ΔresD+abrB were constructed using homologous recombination techniques, and HPLC analyses revealed substantial yield enhancements: 1.6-fold for ΔresD, 2.4-fold for ΔabrB, and a striking 5.8-fold for ΔresD+abrB mutants. In addition, the RT-qPCR results further confirmed that ResD and AbrB were indeed negative regulatory factors for locillomycin. Our results gave rise to a better understanding of the biosynthetic regulation of locillomycin and provided theoretical guidance for the construction of high yielding strains of lipopeptides.

Locillomycin是由velezensis Bs916产生的一种环状脂肽，具有较强的抗菌活性和较低的溶血活性。然而，其产量低已成为大规模应用的重大障碍。为了确定红霉素的调控因子并通过基因敲除获得其高产菌株，我们对loc基因簇的转录格局进行了阐明，并确定了ResD和AbrB是红霉素生物合成基因簇的潜在转录调控因子。EMSA和DNaseⅠ足迹分析结果表明，AbrB倾向于与富含A + t的片段结合，并表现出广泛的dna结合特性，能够直接与loc基因簇的启动子、5' UTR和编码区结合。相比之下，ResD的结合仅限于loc转录起始位点上游的启动子序列，从-480到-456。为了验证它们对红霉素生产的影响，利用同源重组技术构建了突变体ΔresD、ΔabrB和ΔresD+abrB， HPLC分析显示产量大幅提高：ΔresD的产量提高了1.6倍，ΔabrB的产量提高了2.4倍，ΔresD+abrB突变体的产量提高了惊人的5.8倍。此外，RT-qPCR结果进一步证实了ResD和AbrB确实是locillomycin的负调控因子。本研究结果为进一步了解红霉素的生物合成调控提供了理论依据，并为构建高产脂肽菌株提供了理论指导。

{"title":"AbrB and ResD negatively regulate locillomycin synthesis in Bacillus velezensis Bs916 by binding to the flanking sequences of transcription start sites","authors":"Xiaohua Wang , HanWen Shen , Peng Zhang , Zhongzheng Ji , Xiaoxia Xu , Faxiang Wan , Xiulian Yin , Qingyun Bai , Liangcheng Du , Jiasong Jiang , Chuping Luo","doi":"10.1016/j.jbiotec.2025.12.010","DOIUrl":"10.1016/j.jbiotec.2025.12.010","url":null,"abstract":"<div><div>Locillomycin, a cyclic lipopeptide produced by <em>Bacillus velezensis Bs916</em>, has been proven to have strong antimicrobial activity and low hemolytic activity. However, its low yield has become a significant obstacle for large-scale application. To identify the regulatory factors of locillomycin and obtain its high-yield strains through gene knockout, the transcriptional landscape of <em>loc</em> gene cluster was elucidated, and ResD and AbrB were identified as potential transcription regulators of the locillomycin biosynthesis gene cluster. EMSA and DNase Ⅰ footprinting results showed that AbrB tended to bind to A + T-rich fragments and exhibited extensive DNA-binding properties, being able to directly associate with the promoter, 5′ UTR, and coding region of the <em>loc</em> gene cluster. In contrast, the binding of ResD was confined to the promoter sequence upstream of the l<em>oc</em> transcription start site, spanning from −480 to −456. To verify their impact on locillomycin production, mutants <em>ΔresD</em>, <em>ΔabrB</em> and <em>ΔresD+abrB</em> were constructed using homologous recombination techniques, and HPLC analyses revealed substantial yield enhancements: 1.6-fold for <em>ΔresD</em>, 2.4-fold for <em>ΔabrB</em>, and a striking 5.8-fold for <em>ΔresD+abrB</em> mutants. In addition, the RT-qPCR results further confirmed that ResD and AbrB were indeed negative regulatory factors for locillomycin. Our results gave rise to a better understanding of the biosynthetic regulation of locillomycin and provided theoretical guidance for the construction of high yielding strains of lipopeptides.</div></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"410 ","pages":"Pages 207-216"},"PeriodicalIF":3.9,"publicationDate":"2025-12-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145781358","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Semi-rational design of a thermostable O-glycosyltransferase from Glycyrrhiza uralensis for efficient conversion of protopanaxadiol 甘草o -糖基转移酶的半合理设计，用于高效转化原嘌呤二醇。

IF 3.9 2区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Journal of biotechnology

Pub Date : 2025-12-16 DOI: 10.1016/j.jbiotec.2025.12.008

Jiajie Ding, Zhiying Ding, Yuan Liao, Jiajie Chen, Kai Chen, Honghua Jia, Yan Li

Ginsenoside Rh2 (3-β-O-Glc-protopanaxadiol) and Rg3 (20-β-O-Glc-3-β-O-Glc-protopanaxadiol) are rare tetracyclic triterpenoids known for their significant medicinal properties, including antitumor and anti-inflammatory activities. The glycosylation of the C3-OH and C20-OH groups of protopanaxadiol (PPD) by UDP-glycosyltransferase (UGT) is a crucial biological modification that contributes to the extensive structural and functional diversity of PPD-type ginsenosides. In this study, we identified a 3-O-glycosyltransferase (GuUGT73F15) from Glycyrrhiza uralensis, which specifically glycosylates the C3-OH of PPD to produce ginsenoside Rh2. Through site-directed mutagenesis and combinatorial active-site engineering, an optimized variant, GuUGT73F15_{H47P/R84K/N211T}, exhibiting a 2.65-fold increase in catalytic efficiency (k_cat/K_m) for PPD conversion and a remarkable 26.7-fold extension in half-life (t_1/2 = 292.47 min at 37 °C) compared to the wild-type enzyme. Molecular docking and dynamics simulations provided mechanistic insights into the enhanced activity and thermostability of the enzyme. Furthermore, this variant was combined with a glycosyltransferase variant (UGT29_{R91M/D184M/A287V/A342L}) and sucrose synthases, resulting in a high-level production of Rg3 (19.88 mM, 15.61 g/L) in a 24-h fed-batch reaction using PPD as the substrate. This study presents an efficient and thermostable O-glycosyltransferase for the targeted biosynthesis of PPD-type ginsenosides.

人参皂苷Rh2 （3-β-O-Glc-protopanaxadiol）和Rg3 （20-β-O-Glc-3-β-O-Glc-protopanaxadiol）是罕见的四环三萜，具有重要的抗肿瘤和抗炎活性。原人参二醇（PPD）的C3-OH和C20-OH基团被udp -糖基转移酶（UGT）糖基化是PPD型人参皂苷广泛的结构和功能多样性的关键生物学修饰。在本研究中，我们从甘草中鉴定了一种3- o -糖基转移酶（GuUGT73F15），该酶特异性地将PPD的C3-OH糖基化以产生人参皂苷Rh2。通过位点定向诱变和组合活性位点工程，优化后的变体GuUGT73F15H47P/R84K/N211T对PPD转化的催化效率（kcat/Km）比野生型提高了2.65倍，半衰期延长了26.7倍（t1/2 = 292.47min, 37℃）。分子对接和动力学模拟为增强酶的活性和热稳定性提供了机制见解。此外，该变体与糖基转移酶变体（UGT29R91M/D184M/A287V/A342L）和蔗糖合酶偶联，以PPD为底物，在24小时的间歇加料反应中获得了高水平的Rg3 （19.88mM, 15.61g/L）。本研究提出了一种高效、耐热的o -糖基转移酶，用于ppd型人参皂苷的靶向生物合成。

{"title":"Semi-rational design of a thermostable O-glycosyltransferase from Glycyrrhiza uralensis for efficient conversion of protopanaxadiol","authors":"Jiajie Ding, Zhiying Ding, Yuan Liao, Jiajie Chen, Kai Chen, Honghua Jia, Yan Li","doi":"10.1016/j.jbiotec.2025.12.008","DOIUrl":"10.1016/j.jbiotec.2025.12.008","url":null,"abstract":"<div><div>Ginsenoside Rh2 (3-β-O-Glc-protopanaxadiol) and Rg3 (20-β-O-Glc-3-β-O-Glc-protopanaxadiol) are rare tetracyclic triterpenoids known for their significant medicinal properties, including antitumor and anti-inflammatory activities. The glycosylation of the C3-OH and C20-OH groups of protopanaxadiol (PPD) by UDP-glycosyltransferase (UGT) is a crucial biological modification that contributes to the extensive structural and functional diversity of PPD-type ginsenosides. In this study, we identified a 3-O-glycosyltransferase (GuUGT73F15) from <em>Glycyrrhiza uralensis</em>, which specifically glycosylates the C3-OH of PPD to produce ginsenoside Rh2. Through site-directed mutagenesis and combinatorial active-site engineering, an optimized variant, GuUGT73F15<sub>H47P/R84K/N211T</sub>, exhibiting a 2.65-fold increase in catalytic efficiency (<em>k</em><sub>cat</sub>/<em>K</em><sub>m</sub>) for PPD conversion and a remarkable 26.7-fold extension in half-life (t<sub><em>1/2</em></sub> = 292.47 min at 37 °C) compared to the wild-type enzyme. Molecular docking and dynamics simulations provided mechanistic insights into the enhanced activity and thermostability of the enzyme. Furthermore, this variant was combined with a glycosyltransferase variant (UGT29<sub>R91M/D184M/A287V/A342L</sub>) and sucrose synthases, resulting in a high-level production of Rg3 (19.88 mM, 15.61 g/L) in a 24-h fed-batch reaction using PPD as the substrate. This study presents an efficient and thermostable O-glycosyltransferase for the targeted biosynthesis of PPD-type ginsenosides.</div></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"410 ","pages":"Pages 217-227"},"PeriodicalIF":3.9,"publicationDate":"2025-12-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145781418","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Cell specific nutrient feeding in perfusion mode enhances hMSC growth in stirred tank bioreactor process 在搅拌槽生物反应器中，灌注模式下细胞特异性营养物的饲喂可促进hMSC的生长。

IF 3.9 2区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Journal of biotechnology

Pub Date : 2025-12-15 DOI: 10.1016/j.jbiotec.2025.12.006

Johanna Pechan , Kristina Engström , Nora Mac Key , Lina Sörvik , Véronique Chotteau

Human Mesenchymal Stromal Cells (hMSCs) are a safe option for allogeneic cell therapy across various diseases, but their manufacturing process requires improvement to broaden accessibility. In this study, a state-of-the-art planar multi-vessel process was transferred to a stirred tank bioreactor using microcarriers to support the growth of adherent cells. The frequent medium exchange strategy from planar culture guided the design of the bioreactor process. However, complete medium changes in the bioreactor resulted in limited cell expansion and higher glucose consumption compared to planar culture. To enhance expansion, a 0.1 L perfusion bioreactor was tested, enabling continuous medium exchange. Three perfusion approaches were evaluated: (1) maintaining a target cell-specific glucose consumption rate, (2) varying the perfusion rate, and (3) applying a cell-specific perfusion rate. Implementing targeted glucose feeding (TAFE) reduced lactate production, while increasing perfusion rates improved cell density. The highest expansion was achieved using a cell-specific perfusion rate of 5 nL cell⁻¹ day⁻¹ combined with a target glucose consumption rate (q_glc) of 15 pmol cell⁻¹ day⁻¹, resulting in a 5.4-fold higher expansion factor than daily medium changes in stirred tank bioreactors. This optimized process represents a key advancement toward producing clinically relevant quantities of hMSCs.

人间充质基质细胞（hMSCs）是异体细胞治疗多种疾病的安全选择，但其制造工艺需要改进以扩大可及性。在本研究中，将最先进的平面多容器工艺转移到搅拌槽生物反应器中，使用微载体支持贴壁细胞的生长。平面培养的频繁介质交换策略指导了生物反应器工艺的设计。然而，与平面培养相比，生物反应器中完全改变培养基导致细胞扩增有限，葡萄糖消耗更高。为了增强膨胀，我们测试了0.1L的灌注生物反应器，以实现连续的介质交换。评估了三种灌注方法：(1)维持靶细胞特异性葡萄糖消耗率，(2)改变灌注率，(3)应用细胞特异性灌注率。实施靶向葡萄糖喂养（TAFE）减少乳酸生成，而增加灌注率提高细胞密度。当细胞特异性灌注率为5 nL细胞-1天-1，目标葡萄糖消耗率（qglc）为15 pmol细胞-1天-1时，达到了最高的扩增率，其扩增系数比搅拌槽生物反应器中每日培养基变化高5.4倍。这个优化的过程代表了生产临床相关数量的hMSCs的关键进展。

{"title":"Cell specific nutrient feeding in perfusion mode enhances hMSC growth in stirred tank bioreactor process","authors":"Johanna Pechan , Kristina Engström , Nora Mac Key , Lina Sörvik , Véronique Chotteau","doi":"10.1016/j.jbiotec.2025.12.006","DOIUrl":"10.1016/j.jbiotec.2025.12.006","url":null,"abstract":"<div><div>Human Mesenchymal Stromal Cells (hMSCs) are a safe option for allogeneic cell therapy across various diseases, but their manufacturing process requires improvement to broaden accessibility. In this study, a state-of-the-art planar multi-vessel process was transferred to a stirred tank bioreactor using microcarriers to support the growth of adherent cells. The frequent medium exchange strategy from planar culture guided the design of the bioreactor process. However, complete medium changes in the bioreactor resulted in limited cell expansion and higher glucose consumption compared to planar culture. To enhance expansion, a 0.1 L perfusion bioreactor was tested, enabling continuous medium exchange. Three perfusion approaches were evaluated: (1) maintaining a target cell-specific glucose consumption rate, (2) varying the perfusion rate, and (3) applying a cell-specific perfusion rate. Implementing targeted glucose feeding (TAFE) reduced lactate production, while increasing perfusion rates improved cell density. The highest expansion was achieved using a cell-specific perfusion rate of 5 nL cell<sup>−1</sup> day<sup>−1</sup> combined with a target glucose consumption rate (q<sub>glc</sub>) of 15 pmol cell<sup>−1</sup> day<sup>−1</sup>, resulting in a 5.4-fold higher expansion factor than daily medium changes in stirred tank bioreactors. This optimized process represents a key advancement toward producing clinically relevant quantities of hMSCs.</div></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"410 ","pages":"Pages 194-206"},"PeriodicalIF":3.9,"publicationDate":"2025-12-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145774742","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Enhancing oxidative stress defense to improve docosahexaenoic acid production in Schizochytrium sp. HX-308 增强氧化应激防御提高Schizochytrium sp. HX-308二十二碳六烯酸产量

IF 3.9 2区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Journal of biotechnology

Pub Date : 2025-12-12 DOI: 10.1016/j.jbiotec.2025.12.005

Chunxiao Yan , Shuai Zhang , Zhihao Huang , Fei Du , Qian Xu , Wang Ma , Guang Yang , Xiaoman Sun

As an excellent host for docosahexaenoic acid (DHA) biosynthesis, Schizochytrium sp. still faces inherent limitations in DHA yield due to oxidative damage induced by reactive oxygen species (ROS). To effectively reduce the ROS levels, the optimal combination of mixed antioxidants (52.95 mg L⁻¹ phytic acid, 32.00 mg L⁻¹ licorice extract, 36.28 mg L⁻¹ ascorbic acid palmitate) was selected by single factor experiment combined with response surface optimization. The DHA titer increased from 24.0 g L⁻¹ to 33.5 g L⁻¹. However, exogenous addition of antioxidants results in increased the cost of DHA production. Enhancing the intracellular antioxidant capacity through gene engineering has become another feasible strategy. Transcriptome analysis indicated that the transcriptional levels of antioxidant-related genes SOD1 (superoxide dismutase), APX5 (ascorbic acid peroxidase), and GST3 (glutathione S-transferase) were significantly upregulated during DHA biosynthesis. Then, the engineered strain SOD1-APX5-GST3–2 was constructed by overexpressing the three genes, and the DHA and lipid titers were 33.1 g L⁻¹ and 64.1 g L⁻¹, respectively. Scale-up in a 5 L bioreactor obtained 32.5 g L⁻¹ DHA and 63.5 g L⁻¹ lipid, and the ROS level decreased by 30.5 %, demonstrating engineered strain's stability and laying the foundation for industrial-scale production. This study verified that reducing ROS levels promoted DHA biosynthesis in Schizochytrium sp. from the two dimensions of exogenous antioxidants and gene engineering, which provided novel insights for industrial lipid production.

Schizochytrium sp.作为二十二碳六烯酸（二十二碳六烯酸，DHA）生物合成的优良宿主，由于活性氧（reactive oxygen species， ROS）诱导的氧化损伤，其DHA产量仍存在固有的局限性。为有效降低ROS水平，通过单因素试验结合响应面优化，筛选出混合抗氧化剂（52.95mg -1植酸、32.00mg -1甘草提取物、36.28mg -1抗坏血酸棕榈酸酯）的最佳组合。DHA滴度由24.0gL-1提高到33.5gL-1。然而，外源添加抗氧化剂会导致DHA生产成本的增加。通过基因工程增强细胞内抗氧化能力已成为另一种可行的策略。转录组分析表明，抗氧化相关基因SOD1（超氧化物歧化酶）、APX5（抗坏血酸过氧化物酶）和GST3（谷胱甘肽s -转移酶）的转录水平在DHA生物合成过程中显著上调。通过过表达这3个基因构建工程菌株SOD1-APX5-GST3-2，其DHA滴度为33.1gL-1，脂质滴度为64.1gL-1。在5L生物反应器中放大后，得到了32.5gL-1 DHA和63.5gL-1脂质，ROS水平下降了30.5%，证明了工程菌株的稳定性，为工业规模生产奠定了基础。本研究从外源抗氧化剂和基因工程两个维度验证了ROS水平的降低促进了Schizochytrium sp. DHA的生物合成，为工业脂质生产提供了新的见解。

{"title":"Enhancing oxidative stress defense to improve docosahexaenoic acid production in Schizochytrium sp. HX-308","authors":"Chunxiao Yan , Shuai Zhang , Zhihao Huang , Fei Du , Qian Xu , Wang Ma , Guang Yang , Xiaoman Sun","doi":"10.1016/j.jbiotec.2025.12.005","DOIUrl":"10.1016/j.jbiotec.2025.12.005","url":null,"abstract":"<div><div>As an excellent host for docosahexaenoic acid (DHA) biosynthesis, <em>Schizochytrium</em> sp. still faces inherent limitations in DHA yield due to oxidative damage induced by reactive oxygen species (ROS). To effectively reduce the ROS levels, the optimal combination of mixed antioxidants (52.95 mg L<sup>−1</sup> phytic acid, 32.00 mg L<sup>−1</sup> licorice extract, 36.28 mg L<sup>−1</sup> ascorbic acid palmitate) was selected by single factor experiment combined with response surface optimization. The DHA titer increased from 24.0 g L<sup>−1</sup> to 33.5 g L<sup>−1</sup>. However, exogenous addition of antioxidants results in increased the cost of DHA production. Enhancing the intracellular antioxidant capacity through gene engineering has become another feasible strategy. Transcriptome analysis indicated that the transcriptional levels of antioxidant-related genes <em>SOD1</em> (superoxide dismutase), <em>APX5</em> (ascorbic acid peroxidase), and <em>GST3</em> (glutathione S-transferase) were significantly upregulated during DHA biosynthesis. Then, the engineered strain SOD1-APX5-GST3–2 was constructed by overexpressing the three genes, and the DHA and lipid titers were 33.1 g L<sup>−1</sup> and 64.1 g L<sup>−1</sup>, respectively. Scale-up in a 5 L bioreactor obtained 32.5 g L<sup>−1</sup> DHA and 63.5 g L<sup>−1</sup> lipid, and the ROS level decreased by 30.5 %, demonstrating engineered strain's stability and laying the foundation for industrial-scale production. This study verified that reducing ROS levels promoted DHA biosynthesis in <em>Schizochytrium</em> sp. from the two dimensions of exogenous antioxidants and gene engineering, which provided novel insights for industrial lipid production.</div></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"410 ","pages":"Pages 162-172"},"PeriodicalIF":3.9,"publicationDate":"2025-12-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145756688","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Next level of p-phenylene diisothiocyanate-based covalent immobilization of β-D-galactosidase: Technical optimization as an application 基于对苯二异硫氰酸酯的β-d-半乳糖苷酶共价固定化的新水平：技术优化应用。

IF 3.9 2区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Journal of biotechnology

Pub Date : 2025-12-10 DOI: 10.1016/j.jbiotec.2025.12.004

Tabea L. Boehme , Bernadette Straub , Anja Oberle , Ursula Eschenhagen , Magnus S. Schmidt

In this study, a continuous lactose hydrolysis process in a fixed-bed reactor was developed using β-galactosidase covalently immobilized on resin beads via 1,4-phenylenediisothiocyanate (PDC) as linker. Process conditions, including temperature, enzyme loading, perfusion speed, and repeated perfusion of the same substrate solution were systematically varied. The highest glucose yields were obtained at 55 °C, with increased yields observed at low perfusion speeds, high enzyme loadings, and upon repeated perfusions. Under optimized cycle perfusion over 72 h, final lactose conversion reached approximately 90 % at 37 °C and 80 % at 22 °C. A hydrolysis process in a fixed-bed reactor was successfully established, although further optimization is required.

本研究以1,4-苯二异硫氰酸酯（PDC）为连接剂，将β-半乳糖苷酶共价固定在树脂珠上，建立了固定床反应器中乳糖连续水解工艺。工艺条件，包括温度，酶载量，灌注速度，重复灌注相同的底物溶液系统地改变。在55°C时获得最高葡萄糖产量，在低灌注速度、高酶负荷和重复灌注时观察到产量增加。在优化循环灌注超过72h的情况下，最终乳糖转化率在37°C时达到约90%，在22°C时达到80%。在固定床反应器中成功建立了水解过程，但需要进一步优化。

引用次数: 0

Antibacterial effects and mechanisms of fisetin-β-cyclodextrin complex under UVA light against Escherichia coli O157:H7 非瑟酮-β-环糊精配合物在UVA光下对大肠杆菌O157:H7的抑菌作用及其机制

IF 3.9 2区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Journal of biotechnology

Pub Date : 2025-12-06 DOI: 10.1016/j.jbiotec.2025.12.003

Tao Liu , Yuzhang Zhu , Jiahui Wang , Kun Lin , Xuan Zhang , Huawei Ma , Chaonan Kong , Wei Cai , Lifang Yang , Mingguo Jiang

The development of natural product photosensitizers was a critical cornerstone for applying photodynamic antimicrobial technology in the food industry. The objective of this study was to fabricate complex of fisetin-β-cyclodextrin (FIT-β-CD), explore their photodynamic antimicrobial efficacy and mechanisms, and characterize it through infrared spectroscopy and X-ray diffraction. The results indicatde that this method can produce good dispersibility and water solubility of FIT-β-CD complex.The photodynamic impact of FIT-β-CD was assessed by examining alterations in reactive oxygen species (ROS) production.The results demonstrated that FIT-β-CD can rapidly generate ROS under UVA, with hydrogen peroxide production being 3.73 times higher than in dark conditions. The antimicrobial efficacy of FIT-β-CD against Escherichia coli O157:H7 (E.coli O157:H7) was evaluated based on bactericidal efficacy, cell membrane damage, DNA and protein damage, motility changes, biofilm formation, and extracellular polysaccharide production. In addition, gene expression analysis by RT-qPCR showed its significant inhibition on the two-component system (TCS) and quorum sensing system (QS). The results indicated that FIT-β-CD exhibited efficient, sensitive photodynamic, and antibacterial properties against E.coli O157:H7, making it a novel photosensitizer. These findings imply potential new applications of fisetin in the field of food antimicrobial agents.

天然产物光敏剂的开发是光动力抗菌技术在食品工业中应用的重要基石。本研究的目的是制备非瑟酮-β-环糊精配合物（FIT-β-CD），探讨其光动力抗菌效果和机制，并通过红外光谱和x射线衍射对其进行表征。结果表明，该方法制备的FIT-β-CD配合物具有良好的分散性和水溶性。通过检测活性氧（ROS）产生的变化来评估FIT-β-CD的光动力学影响。结果表明，在UVA条件下，FIT-β-CD可以快速生成ROS，过氧化氢产量是暗条件下的3.73倍。通过对大肠杆菌O157:H7 （E.coli O157:H7）的杀菌效果、细胞膜损伤、DNA和蛋白质损伤、运动改变、生物膜形成和胞外多糖生成等指标评价FIT-β-CD对大肠杆菌O157:H7的抑菌效果。此外，RT-qPCR基因表达分析显示其对双组分系统（TCS）和群体感应系统（QS）有显著抑制作用。结果表明，FIT-β-CD对大肠杆菌O157:H7具有高效、灵敏的光动力学和抗菌性能，是一种新型光敏剂。这些发现暗示了非瑟酮在食品抗菌剂领域的潜在新应用。

{"title":"Antibacterial effects and mechanisms of fisetin-β-cyclodextrin complex under UVA light against Escherichia coli O157:H7","authors":"Tao Liu , Yuzhang Zhu , Jiahui Wang , Kun Lin , Xuan Zhang , Huawei Ma , Chaonan Kong , Wei Cai , Lifang Yang , Mingguo Jiang","doi":"10.1016/j.jbiotec.2025.12.003","DOIUrl":"10.1016/j.jbiotec.2025.12.003","url":null,"abstract":"<div><div>The development of natural product photosensitizers was a critical cornerstone for applying photodynamic antimicrobial technology in the food industry. The objective of this study was to fabricate complex of fisetin-β-cyclodextrin (FIT-β-CD), explore their photodynamic antimicrobial efficacy and mechanisms, and characterize it through infrared spectroscopy and X-ray diffraction. The results indicatde that this method can produce good dispersibility and water solubility of FIT-β-CD complex.The photodynamic impact of FIT-β-CD was assessed by examining alterations in reactive oxygen species (ROS) production.The results demonstrated that FIT-β-CD can rapidly generate ROS under UVA, with hydrogen peroxide production being 3.73 times higher than in dark conditions. The antimicrobial efficacy of FIT-β-CD against <em>Escherichia coli</em> O157:H7 (<em>E.coli</em> O157:H7) was evaluated based on bactericidal efficacy, cell membrane damage, DNA and protein damage, motility changes, biofilm formation, and extracellular polysaccharide production. In addition, gene expression analysis by RT-qPCR showed its significant inhibition on the two-component system (TCS) and quorum sensing system (QS). The results indicated that FIT-β-CD exhibited efficient, sensitive photodynamic, and antibacterial properties against <em>E.coli</em> O157:H7, making it a novel photosensitizer. These findings imply potential new applications of fisetin in the field of food antimicrobial agents.</div></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"410 ","pages":"Pages 145-152"},"PeriodicalIF":3.9,"publicationDate":"2025-12-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145708122","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Increasing the chiral purity of cellulosic L-lactic acid by D-Lactate oxidase-catalyzed oxidation in engineered lactic acid bacterium Pediococcus acidilactici d -乳酸氧化酶在工程乳酸菌中催化氧化提高纤维素l -乳酸的手性纯度。

IF 3.9 2区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Journal of biotechnology

Pub Date : 2025-12-05 DOI: 10.1016/j.jbiotec.2025.12.002

Jiao Liu, Chaolong Qu, Bin Zhang, Jie Bao

Trace amounts of D-lactic acid derived from lignocellulose feedstock and nitrogen additives frequently reduce the chiral purity of L-lactic acid below the polymerization standard during biorefinery fermentations. The spontaneous D-lactic acid degradation catalyzed by the highly substrate specific D-lactate oxidase in lactic acid bacterium provides an ideal solution. This study used a D-lactate oxidase GOX2071 with relatively high activity for construction of D-lactic acid oxidation pathway in the L-lactic acid producing strain Pediococcus acidilactici ZY271. The engineered P. acidilactici LJ2071 strain demonstrated an active D-lactic acid oxidation performance while the high L-lactic acid production property was well maintained. A L-lactic acid chiral purity of 99.63 % was obtained using the D-lactic acid containing wheat straw as feedstock and 40 % of corn steep liquor (CSL) as nitrogen additive to alternate expensive yeast extract. This study provided a practical method for upgrading the chirality of cellulosic L-lactic acid using lignocellulose feedstock and cheap nitrogen additives.

在生物精炼发酵过程中，来自木质纤维素原料和氮添加剂的微量d -乳酸经常使l -乳酸的手性纯度低于聚合标准。乳酸菌中具有高度底物特异性的d -乳酸氧化酶对d -乳酸的自发降解提供了理想的解决方案。本研究利用活性较高的d -乳酸氧化酶GOX2071在产l -乳酸的Pediococcus acidacactii ZY271中构建d -乳酸氧化途径。工程菌株LJ2071在保持高产l -乳酸特性的同时，表现出较强的d -乳酸氧化能力。以小麦秸秆为原料，以40%的玉米浸泡液（CSL）为氮添加剂，替代昂贵的酵母浸膏，得到了手性纯度为99.63%的l -乳酸。本研究为利用木质纤维素原料和廉价的氮添加剂提高纤维素l -乳酸的手性提供了一种实用的方法。

{"title":"Increasing the chiral purity of cellulosic L-lactic acid by D-Lactate oxidase-catalyzed oxidation in engineered lactic acid bacterium Pediococcus acidilactici","authors":"Jiao Liu, Chaolong Qu, Bin Zhang, Jie Bao","doi":"10.1016/j.jbiotec.2025.12.002","DOIUrl":"10.1016/j.jbiotec.2025.12.002","url":null,"abstract":"<div><div>Trace amounts of <span>D</span>-lactic acid derived from lignocellulose feedstock and nitrogen additives frequently reduce the chiral purity of <span>L</span>-lactic acid below the polymerization standard during biorefinery fermentations. The spontaneous <span>D</span>-lactic acid degradation catalyzed by the highly substrate specific <span>D</span>-lactate oxidase in lactic acid bacterium provides an ideal solution. This study used a <span>D</span>-lactate oxidase GOX2071 with relatively high activity for construction of <span>D</span>-lactic acid oxidation pathway in the <span>L</span>-lactic acid producing strain <em>Pediococcus acidilactici</em> ZY271. The engineered <em>P</em>. <em>acidilactici</em> LJ2071 strain demonstrated an active <span>D</span>-lactic acid oxidation performance while the high <span>L</span>-lactic acid production property was well maintained. A <span>L</span>-lactic acid chiral purity of 99.63 % was obtained using the <span>D</span>-lactic acid containing wheat straw as feedstock and 40 % of corn steep liquor (CSL) as nitrogen additive to alternate expensive yeast extract. This study provided a practical method for upgrading the chirality of cellulosic <span>L</span>-lactic acid using lignocellulose feedstock and cheap nitrogen additives.</div></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"410 ","pages":"Pages 137-144"},"PeriodicalIF":3.9,"publicationDate":"2025-12-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145701077","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Enhancing heterotrophic lutein production in Chlorella protothecoides through combined phytohormone and nitrogen strategies 植物激素和氮素联合策略促进原小球藻异养叶黄素的产生

IF 3.9 2区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Journal of biotechnology

Pub Date : 2025-12-04 DOI: 10.1016/j.jbiotec.2025.12.001

Jiaxin Li , Weiyang Zhao , Yunlei Fu , Ranxuan Li , Jin Liu , Shufang Yang , Jie Zheng , Haijin Mou , Han Sun

Large-scale production of microalgal lutein faces challenges due to limited biomass and lutein content. In this work, we developed an exponential fed-batch model to achieve heterotrophic high-density growth of Chlorella protothecoides CS-41. Then, indole-3-acetic acid (IAA) and high nitrogen were combined to promote lutein accumulation in situ. Fed-batch culture enhanced the TCA cycle of C. protothecoides CS-41 to benefit cell growth while increasing carotenoid flux to facilitate lutein accumulation through redirecting carbon flux towards lutein biosynthesis, improved by IAA. Also, high nitrogen increased lutein content through modulation of CMS (carotenoid methyltransferase synthase), CRTISO (carotenoid isomerase), LCYB (lycopene β-cyclase), ZEP (zeaxanthin epoxidase), and VDE (violaxanthin de-epoxidase). Finally, under heterotrophic conditions, combining 200 mM IAA with 3.6 g/L urea improved pyruvate and 3-phosphoglyceric acid and then boosted lutein content and productivity to 3.27 mg/g and 20.38 mg/L/d, respectively, by increases of 58 % and 56.7 %. Addressing issues including density dilution, container transfer, and in-situ lutein accumulation under heterotrophic high-density conditions offers a cost-effective solution.

由于生物量和叶黄素含量有限，微藻叶黄素的大规模生产面临挑战。本研究建立了小球藻CS-41异养高密度生长的指数补料批量模型。然后，吲哚-3-乙酸（IAA）和高氮配合施用，促进叶黄素原位积累。补料批培养增强了C. protothecoides CS-41的TCA循环，有利于细胞生长，同时增加类胡萝卜素通量，通过将碳通量转向叶黄素的生物合成，促进叶黄素的积累，IAA改善了这一过程。高氮通过调节类胡萝卜素甲基转移酶合成酶（CMS）、类胡萝卜素异构酶（CRTISO）、番茄红素β环化酶（LCYB）、玉米黄质环氧化酶（ZEP）和紫黄质去环氧化酶（VDE）提高叶黄素含量。最后，在异养条件下，200 mM IAA与3.6 g/L尿素配用可改善丙酮酸和3-磷酸甘油酸，叶黄素含量和产量分别提高了3.27 mg/g和20.38 mg/L/d，分别提高了58 %和56.7% %。在异养高密度条件下，解决密度稀释、容器转移和原位叶黄素积累等问题提供了具有成本效益的解决方案。

{"title":"Enhancing heterotrophic lutein production in Chlorella protothecoides through combined phytohormone and nitrogen strategies","authors":"Jiaxin Li , Weiyang Zhao , Yunlei Fu , Ranxuan Li , Jin Liu , Shufang Yang , Jie Zheng , Haijin Mou , Han Sun","doi":"10.1016/j.jbiotec.2025.12.001","DOIUrl":"10.1016/j.jbiotec.2025.12.001","url":null,"abstract":"<div><div>Large-scale production of microalgal lutein faces challenges due to limited biomass and lutein content. In this work, we developed an exponential fed-batch model to achieve heterotrophic high-density growth of <em>Chlorella protothecoides</em> CS-41. Then, indole-3-acetic acid (IAA) and high nitrogen were combined to promote lutein accumulation <em>in situ</em>. Fed-batch culture enhanced the TCA cycle of <em>C. protothecoides</em> CS-41 to benefit cell growth while increasing carotenoid flux to facilitate lutein accumulation through redirecting carbon flux towards lutein biosynthesis, improved by IAA. Also, high nitrogen increased lutein content through modulation of <em>CMS</em> (carotenoid methyltransferase synthase), <em>CRTISO</em> (carotenoid isomerase), <em>LCYB</em> (lycopene β-cyclase), <em>ZEP</em> (zeaxanthin epoxidase), and <em>VDE</em> (violaxanthin de-epoxidase). Finally, under heterotrophic conditions, combining 200 mM IAA with 3.6 g/L urea improved pyruvate and 3-phosphoglyceric acid and then boosted lutein content and productivity to 3.27 mg/g and 20.38 mg/L/d, respectively, by increases of 58 % and 56.7 %. Addressing issues including density dilution, container transfer, and <em>in-situ</em> lutein accumulation under heterotrophic high-density conditions offers a cost-effective solution.</div></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"410 ","pages":"Pages 125-136"},"PeriodicalIF":3.9,"publicationDate":"2025-12-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145681983","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

High specificity of MsmCAR toward 4-hydroxyvaleric acid enables efficient 1,4-pentanediol production from biomass-derived levulinic acid MsmCAR对4-羟戊酸的高特异性使得从生物质衍生的乙酰丙酸中高效生产1,4-戊二醇。

IF 3.9 2区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Journal of biotechnology

Pub Date : 2025-12-01 DOI: 10.1016/j.jbiotec.2025.11.020

Suhye Choi , Seung Hun Lee , Yunyeong Kim , Woo Young Jeon , Jung oh Ahn , Shashi Kant Bhatia , Kwon-Young Choi , Jeong Chan Joo , Yung-Hun Yang

Alkanediols are industrial compounds with diverse applications. However, the biological production of C5 diols is limited by the low substrate specificity and efficiency of carboxylic acid reductases (CARs) toward short to medium-chain substrates. We developed a whole-cell biotransformation system for efficient 1,4-pentanediol (1,4-PDO) production, aimed at developing a platform pathway for diol biosynthesis. By screening 10 CARs, we identified MsmCAR from Mycolicibacterium smegmatis MC2 155 as the most effective for C5 diol biosynthesis, with high specificity toward 4-hydroxyvaleric acid (4-HV). We constructed a whole-cell catalyst expressing MsmCAR and enhanced its performance by introducing ppk2b from Corynebacterium glutamicum ATCC 13032 and chnD from Acinetobacter sp., which encode proteins involved in cofactor regeneration. Systematic optimization of the reaction conditions including buffers, cofactors, metal ions, and cultivation parameters led to a maximum titer of 78.10 mM 1,4-PDO, with a productivity of 1.86 mM/h, when using 4-HV as a substrate. To expand the substrate scope to levulinic acid, 3-hydroxybutyrate dehydrogenase and formate dehydrogenase biosynthetic genes were introduced. This system yielded 22.10 mM 1,4-PDO within 10 h. This work highlights the potential of MsmCAR as a versatile biocatalyst and provides a sustainable strategy for producing short- to medium-chain diols from biomass-derived feedstocks.

烷二醇是一种用途广泛的工业化合物。然而，C5二醇的生物生产受到低底物特异性和羧酸还原酶（CARs）对中短链底物的效率的限制。我们开发了一种高效生产1,4-戊二醇（1,4- pdo）的全细胞生物转化系统，旨在开发二醇生物合成的平台途径。通过筛选10个CARs，我们鉴定出来自垢垢分枝杆菌MC2 155的MsmCAR对C5二醇的生物合成最有效，对4-羟基戊酸（4-HV）具有高特异性。我们构建了表达MsmCAR的全细胞催化剂，并通过引入来自谷氨棒状杆菌ATCC 13032的ppk2b和来自不动杆菌sp.的chnD来增强其性能，这两个基因编码参与辅助因子再生的蛋白质。系统优化反应条件，包括缓冲液、辅因子、金属离子和培养参数，以4-HV为底物时，最大滴度为78.10mM 1,4- pdo，产率为1.86mM/h。为了将底物范围扩大到乙酰丙酸，引入了3-羟基丁酸脱氢酶和甲酸脱氢酶生物合成基因。该体系在10h内生成22.10mM的1,4- pdo。这项工作突出了MsmCAR作为一种多功能生物催化剂的潜力，并为从生物质来源的原料中生产短至中链二醇提供了可持续的策略。

{"title":"High specificity of MsmCAR toward 4-hydroxyvaleric acid enables efficient 1,4-pentanediol production from biomass-derived levulinic acid","authors":"Suhye Choi , Seung Hun Lee , Yunyeong Kim , Woo Young Jeon , Jung oh Ahn , Shashi Kant Bhatia , Kwon-Young Choi , Jeong Chan Joo , Yung-Hun Yang","doi":"10.1016/j.jbiotec.2025.11.020","DOIUrl":"10.1016/j.jbiotec.2025.11.020","url":null,"abstract":"<div><div>Alkanediols are industrial compounds with diverse applications. However, the biological production of C5 diols is limited by the low substrate specificity and efficiency of carboxylic acid reductases (CARs) toward short to medium-chain substrates. We developed a whole-cell biotransformation system for efficient 1,4-pentanediol (1,4-PDO) production, aimed at developing a platform pathway for diol biosynthesis. By screening 10 CARs, we identified <em>Msm</em>CAR from <em>Mycolicibacterium smegmatis</em> MC2 155 as the most effective for C5 diol biosynthesis, with high specificity toward 4-hydroxyvaleric acid (4-HV). We constructed a whole-cell catalyst expressing <em>Msm</em>CAR and enhanced its performance by introducing <em>ppk2b</em> from <em>Corynebacterium glutamicum</em> ATCC 13032 and <em>chnD</em> from <em>Acinetobacter</em> sp., which encode proteins involved in cofactor regeneration. Systematic optimization of the reaction conditions including buffers, cofactors, metal ions, and cultivation parameters led to a maximum titer of 78.10 mM 1,4-PDO, with a productivity of 1.86 mM/h, when using 4-HV as a substrate. To expand the substrate scope to levulinic acid, 3-hydroxybutyrate dehydrogenase and formate dehydrogenase biosynthetic genes were introduced. This system yielded 22.10 mM 1,4-PDO within 10 h. This work highlights the potential of <em>Msm</em>CAR as a versatile biocatalyst and provides a sustainable strategy for producing short- to medium-chain diols from biomass-derived feedstocks.</div></div>","PeriodicalId":15153,"journal":{"name":"Journal of biotechnology","volume":"410 ","pages":"Pages 115-124"},"PeriodicalIF":3.9,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145668412","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0