ACS Synthetic Biology
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2025
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IF 3.9
Zheng Lei,Jinyong Wu,Caiwen Lao,Jin Wang,Yanyi Xu,He Li,Lixia Yuan,Xiangsong Chen,Jianming Yao
Ectoine is an important natural macromolecule protector that helps extremophiles maintain cellular stability and function under high-salinity conditions. Recently, the development of microbial strains for high-level ectoine production has become an attractive research direction. In this study, we constructed an efficient plasmid-free ectoine-producing strain. We modified the 5′-untranslated region of the ectABC gene cluster from Halomonas elongate to fine-tune the expression of genes ectA, ectB, and ectC. Furthermore, we optimized the carbon flow across the MEP pathway, the TCA cycle, and the aspartic acid metabolic pathway. Subsequently, we blocked the production of byproducts from the aspartic acid metabolic pathway and dynamically regulated the TCA cycle to coordinate the balance between strain growth and production. The final strain was tested in a 5-L fermenter, which reached 118.5 g/L at 114 h of fermentation. The metabolic engineering strategies employed in this study can be used for the biosynthesis of other aspartate derivatives.
研究领域:
微生物学
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生物技术
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PubMed
高影响因子
ACS Synthetic Biology
·
2025
·
IF 3.9
Yifan Feng,Wenlong Xiao,Xinyi Li,Weiyu Cao,Yujia Jiang,Wenming Zhang,Wankui Jiang,Fengxue Xin,Min Jiang
Ectoine, a major compatible solute in halophilic micro-organisms, shows great potential in cosmetics and pharmaceuticals areas owing to its water-binding properties and capability to prevent oxidative damage. In this study, the ectABC gene cluster responsible for the ectoine synthesis originated from halophilic bacterium Halomonas venusta was first assembled into Escherichia coli. Subsequently, the crr gene in PTS was knocked out to further drive the metabolic flux from phosphoenolpyruvate to oxaloacetate, resulting in 1.27 g/L of ectoine. Then, the rate-limiting enzyme LysC in the ectoine synthesis pathway was identified and modified. The recombinant E. coli with the further overexpression of feedback-insensitive mutant EclysC* increased the ectoine titer to 2.51 g/L with a yield of 0.37 g/g in shake flasks. After the medium optimization including the carbon and nitrogen source, sodium chloride, and magnesium sulfate concentration, the ectoine titer was improved to 4.55 g/L. 115.15 g/L of ectoine with a yield of 0.23 g/g was obtained in the 5.0 L bioreactor through the optimization of substrate feeding and IPTG supplementation in the fed-batch fermentation. To achieve the cost-effective production of ectoine, lignocellulosic hydrolysate from wheat straw was adopted. 134.08 g/L of ectoine with a yield of 0.33 g/g sugar and a productivity of 3.7 g/L/h was finally produced, representing a relatively high level of ectoine production from renewable resources compared to other studies. This study provides valuable insights into a cost-effective and efficient method for industrial-scale ectoine production.
研究领域:
生物技术
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微生物学
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PubMed
高影响因子
INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
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2024
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IF 7.7
Bingmei Su,Wen Yang,Yi Zhou,Juan Lin
Ectoine, so-called tetrahydropyrimidine, is an important osmotic adjustment solute and widely applied in cosmetics and protein protectant. Some attempts have been made to improve the ectoine productivity. However, the strains with both high ectoine production capacity and high glucose conversion were still absent so far. Aim to construct a strain for efficiently producing ectoine, ectoine synthetic gene cluster ectABC from Pseudomonas stutzeri was overexpressed in E. coli BL21 (DE3). The ection production was improved by 382 % (ectoine titer increased from 1.73 g/L to 8.33 g/L) after the rational design of rate-limiting enzyme L-2,4-diaminobutyrate transaminase EctB ps (protein engineering) combined with the metabolic engineering that focused on the enrichment and conversion of precursors. The final strain YW20 was applied to overproduce ectoine in fed-batch fermentation and yield 68.9 g/L of ectoine with 0.88 g/L/h of space-time yield and the highest glucose conversion reported [34 % (g/g)]. From the fermentation broth, ectoine was purified with 99.7 % purity and 79.8 % yield. This study successfully provided an engineered strain as well as an efficient method for the industrial bio-synthesis and preparation of ectoine.
研究领域:
生物技术
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微生物学
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生物化工
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PubMed
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COLLOIDS AND SURFACES A-PHYSICOCHEMICAL AND ENGINEERING ASPECTS
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2025
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IF 5.4
Li Xu,Zhenlin Shang,Dongxu Liu,Caiyun Zhao,Chengke Zhao,Xiaojing Pei,Ze Zhang
In biomedicine and cosmetics, skin-targeted drug delivery is crucial yet challenging due to the skin complex barrier. Low molecular weight hyaluronic acid (HA) shows targeting ability as it can bind to CD44 receptors overexpressed on fibroblasts. Two types of targeted liposomes were developed in this study, covalently modified hyaluronic acid-ectoine liposome (HA-ECT-LPs) and non-covalently modified hyaluronic acid-chitosan-ectoine liposome (HA-CS-ECT-LPs). DSPE can stabilize liposomes through the interaction of low molecular weight HA chains in bilayer, and can be covalently modified on the surface of liposomes after coupling with HA. The non-covalently modified liposomes adsorbed HA electrostatic onto the surface of chitosan-coated ectodoin liposomes, which can easily and quickly form multilayer liposomes and delay drug release. The sizes, PDIs, and zeta potentials particle sizes of HA-ECT-LPs and HA-CS-ECT-LPs were determined to be 280 ± 4 nm and 232 ± 7 nm, PDI were 0.30 ± 0.04 and 0.50 ± 0.11, Zeta potential were -24 ± 2 mV and -13 ± 1 mV, respectively. The 90 days stability tests showed HA-ECT-LPs and HA-CS-ECT-LPs has good stability, ΔBS were less than 0.5%. Drug release test showed that HA-ECT-LPs and HA-CS-ECT-LPs with slower release than ectoine liposomes in vitro . Skin permeation proved that compared with ectoine liposomes, the cumulative release amount of HA-ECT-LPs and HA-CS-ECT-LPs increased by 41% and 31%, and the cumulative retention amount increased by 21% and 10%, respectively. Cell experiments demonstrated that both types of liposomes have excellent targeting to fibroblasts. The evaluation of the efficacy of liposomes on human skin indicated that for the essence liquids of HA-ECT-LPs and HA-CS-ECT-LPs, after 7 days, the skin water content increased by 2.51 times and 2.56 times respectively, and the transepidermal water loss (TEWL) decreased by 1.69 times and 2.45 times respectively. After 30 days, the skin elasticity increased by 2.88 times and 2.29 times respectively. Overall, these liposomes offered great potential for skin delivery of ectoine, offering a promising approach for skin health applications.
研究领域:
生物医学
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化妆品科学
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药剂学
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DOI
高影响因子
ACS Synthetic Biology
·
2025
·
IF 3.9
Zheng Lei,Xiangsong Chen,Lixia Yuan,Jinyong Wu,Jianming Yao
Ectoine is a pivotal natural osmoprotectant that functions as a compatible solute through osmoregulation, enabling microorganisms to thrive in extreme environments such as high salinity. To meet market demands, this study focuses on optimizing its production process. We initially engineered the ectABC gene cluster from Halomonas venusta via 5′-UTR modification, establishing a functional ectoine biosynthesis pathway in E. coli. Subsequent introduction of a rate-limiting enzyme EctB mutant (E407D) and aspartokinase mutant increased titer by 140%. To address lysine byproduct accumulation, an innovative molecular switch was employed to regulate lysA gene expression, achieving dynamic balance between cell growth and product synthesis. Further optimization through cofactor engineering yielded the final strain ECT31, which produced 164.6 g/L ectoine in a 100 L bioreactor within 117 h, the highest reported titer for E. coli-based ectoine production to date. The metabolic engineering strategy presented herein establishes a new pdigm for efficient biosynthesis of amino acid derivatives.
研究领域:
合成生物学
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生物技术
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微生物学
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生物工程
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分子生物学
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PubMed
高影响因子
ANNALS OF CLINICAL AND LABORATORY SCIENCE
·
2025
·Vol. 55
·期 6
·939-953
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IF 1
Yu Zhang,Guangju Zhou
Objective Ectoine is a natural compatible solute known for its cytoprotective properties, though its potential role in alleviatingdiabetic cardiomyopathy remains largely unexplored. This study aimed to examine the protective effects and underlying mechanismsof Ectoine on H9c2 cardiomyocytes under high-glucose and high-palmitic acid conditions. Methods The investigation included assessment of cell viability and lactate dehydrogenase release. Oxidative stress was evaluatedthrough measurements of reactive oxygen species, malondialdehyde levels, and superoxide dismutase activity. Inflammatory responseswere analyzed via TNF-α secretion and NF-κB and ERK1/2 pathways. Molecular docking studies were conducted to explore Ectoine-AMPK interaction, while mechanistic insightswere gained through examination of AMPK signaling, energy metabolism regulators, autophagic flux markers, and mitochondrialapoptosis. Results Ectoine treatment was associated with enhanced cell viability and reduced lactate dehydrogenase release. It appeared to alleviateoxidative stress through decreased reactive oxygen species and malondialdehyde levels, along with increased superoxide dismutaseactivity. Ectoine also showed suppressive effects on inflammatory responses, including reduced TNF-α secretion and inhibited NF-κB and ERK1/2 pathways. Molecular docking suggested binding affinity between Ectoine and AMPK (−6.4 kcal/mol). Mechanistically,Ectoine demonstrated potential in activating AMPK signaling, modulating energy metabolism, restoring autophagic flux, andattenuating mitochondrial apoptosis. Conclusion The findings suggest that Ectoine may protect against HG-PA-induced cardiomyocyte injury. This protective effect was associatedwith AMPK activation and the coordinated modulation of metabolic, autophagic, anti-inflammatory, and anti-apoptotic pathways.This study suggests the therapeutic potential of Ectoine and provides a basis for further investigation.
研究领域:
心血管研究
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分子生物学
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细胞生物学
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代谢性疾病
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药理学
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PubMed
高影响因子
ACS Synthetic Biology
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2026
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IF 4.5
Yanyan Lei,Yiyao Dong,Songlin Zhang,Huimin Zhang,Rongshuai Zhu,Minglong Shao,Zhiming Rao
Ectoine is a highly valuable amino acid derivative with multiple functionalities, which finds extensive applications in cosmetics, pharmaceuticals, and life sciences. Developing microbial strains capable of high-level ectoine production has emerged as a prominent research focus in recent years. Here, we employed a systematic metabolic engineering strategy to transform wild-type Escherichia coli into a high-yield ectoine-producing strain. First, we constructed a heterologous ectoine synthesis pathway in E. coli W3110. By knocking out the bifunctional enzymes ThrA and MetL, as well as LysA to block byproduct formation and overexpressing an optimized LysC as a substitute, we enhanced the supply of the direct precursor. We also investigated the impact of the copy number on ectoine synthesis. Subsequently, we modified the 5′ untranslated region of citrate dehydrogenase gltA to fine-tune its expression, balancing cellular growth with product synthesis. To augment glutamate amino donor availability, we heterologously overexpressed Bacillus subtilis gltAB to enhance glutamate supply, while boosting pntAB expression to maintain cofactor equilibrium. Similarly, we fortified the glucose-to-oxaloacetate synthetic pathway through a series of metabolic modifications, achieving a yield of 5.94 g/L in shake flask fermentation. Finally, under controlled batch glucose feeding, strain E20 produced 88.1 g/L of ectoine over a 60 h fermentation period, and a glucose conversion rate of 0.26 g/g. This study employed metabolic engineering strategies to enhance the accumulation of oxaloacetate and utilized 5′-UTR engineering to finely regulate GltA expression, thereby balancing cell growth, These strategies, including increasing aspartate accumulation, enhancing the catalytic efficiency of the key heterologous enzyme LysCpa in the aspartate to aspartate phosphate pathway, and boosting glutamate as an amino donor to enhance the synthesis of aspartate from oxaloacetic acid, can be applied to the synthesis of other amino acids in the aspartate family.
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Microbial Cell Factories
·
2024
·Vol. 23
·期 1
·1-12
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IF 4.3
Chen Yaping,Liu Yujie,Meng Yan,Jiang Yuting,Xiong Weini,Wang Shufang,Yang Chao,Liu Ruihua
Background Ectoine as an amino acid derivative is widely applied in many fields, such as the food industry, cosmetic manufacturing, biologics, and therapeutic agent. Large-scale production of ectoine is mainly restricted by the cost of fermentation substrates (e.g., carbon sources) and sterilization. Results In this study, Halomonas cupida J9 was shown to be capable of synthesizing ectoine using xylose as the sole carbon source. A pathway was proposed in H. cupida J9 that synergistically utilizes both WBG xylose metabolism and EMP glucose metabolism for the synthesis of ectoine. Transcriptome analysis indicated that expression of ectoine biosynthesis module was enhanced under salt stress. Ectoine production by H. cupida J9 was enhanced by improving the expression of ectoine biosynthesis module, increasing the intracellular supply of the precursor oxaloacetate, and utilizing urea as the nitrogen source. The constructed J9U-P8EC achieved a record ectoine production of 4.12 g/L after 60 h of xylose fermentation. Finally, unsterile production of ectoine by J9U-P8EC from either a glucose-xylose mixture or corn straw hydrolysate was demonstrated, with an output of 8.55 g/L and 1.30 g/L of ectoine, respectively. Conclusions This study created a promising H. cupida J9-based cell factory for low-cost production of ectoine. Our results highlight the potential of J9U-P8EC to utilize lignocellulose-rich agriculture waste for open production of ectoine.
研究领域:
微生物学
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生物技术
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代谢工程
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DOI
PubMed
高影响因子
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