ANALYTICAL CHEMISTRY
·
2025
·
IF 6.7
Yan Gao,Quanzhi Xiao,Jie Zhang,Kena Zhang,Liping Fang,Xiao-Xia Zhou,Bing Yan
Nanoplastics (NPs) are prevalent in the environment, posing risks to ecosystems and human health. While research into their effects on bacterial activity has increased, the mechanisms underlying NP-bacteria interactions─specifically whether NPs penetrate cells or adhere to the cell surface─remain poorly understood. This knowledge gap largely stems from the absence of quantitative analytical methods. Herein, we developed a novel approach combining lysozyme treatment with pyrolysis gas chromatography–mass spectrometry (Py-GC/MS) to differentiate between intracellular and cell wall-bound NPs in Escherichia coli (E. coli) quantitatively. The method involves selective removal of the bacterial cell wall using lysozyme, protein corona-induced extraction to enrich cell wall-bound NPs, and hydrogen peroxide digestion to eliminate protoplast interference before Py-GC/MS analysis. Validation with europium (Eu)-labeled NPs, quantified by inductively coupled plasma mass spectrometry (ICP-MS), confirmed the method’s accuracy and reliability. Using this approach, we found that after NP exposure, only a small fraction (9.6–10.5%) of NPs penetrated E. coli cells, while the majority (36.9–63.8%) adhered to the cell surface. Transmission electron microscopy further corroborated these findings. Consequently, this work provides a robust tool for the quantification of NP uptake and biodistribution in bacterial systems, advancing our understanding of NP–microorganism interactions and their environmental implications.
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BioDesign Research
·
2026
·
IF 7.2
Haidong Yu,Haofeng Chen,Guojun Yang,Zhen Huang,Jieming Zhou,Dongjiang Lin,Junru Zhao,Yang Zhang,Jifeng Yuan
1-Butanol is a C4 aliphatic alcohol that is widely used as a gasoline blendstock. In this study, we report an underexplored metabolic route for the de novo biosynthesis of 1-butanol in Escherichia coli . Specifically, the 1-butanol biosynthetic pathway comprises a redesigned Clostridium -derived pathway for butyryl-CoA synthesis, followed by a CoA-acylating aldehyde dehydrogenase (PduP from Salmonella enterica ) and an endogenous alcohol dehydrogenase (YqhD from E. coli ). We found that abolishing thioesterases to block the formation of butyrate byproduct would lead to deleterious effects on the cell growth. Upon introducing a metabolic buffering pathway towards polyhydroxyalkanoate biosynthesis, the toxicity caused by metabolic imbalance was alleviated, accompanied by enhanced cell growth and 1-butanol productivity. A similar strategy was also validated to be effective for improving the production of hexane-2,3-diol, a potential cosmetic component with high value. In summary, this work successfully demonstrates that the metabolic buffering strategy exerts a synergistic effect on enhancing biochemical production with improved carbon recovery efficiency.
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Environmental Technology & Innovation
·
2025
·
IF 7.1
Yinhua Cui,Yiwen Chen,Qingshan Shi,Jin Feng,Yafeng Song,Junwei Yao,Xiaobao Xie
A growing number of microorganisms are employed in industry nowadays, leading to the extensive generation of microbial waste. Nevertheless, the waste recycling investigation has not yet encompassed the synthesis of nanoparticles. Economically viable nano-selenium (nano-Se) requires additional cost reduction. This study produced cost-effective nano-Se utilizing microbial waste. This reaction was conducted under UV irradiation due to the low activity of microbial waste. After 90 min of UV irradiation, the yield of nano-Se generated from the death-phase eukaryote Saccharomyces cerevisiae ( S. cerevisiae ) waste was 7.46 times more than that of nano-Se produced in darkness. The resultant nano-Se is amorphous, and the nano-Se generated by UV irradiation demostrated a larger size than that produced in darkness. Moreover, unlike the nano-Se generated in darkness or incubators, the nano-Se produced under UV irradiation significantly inhibited the multiplication of MDA-MB-231 cancer cells, demonstrating its enhanced efficacy. The UV-assisted synthesis of nano-Se was also seen during the death phase of the prokaryote Escherichia coli ( E. coli ) waste. After 5 h of UV irradiation, the yield of nano-Se produced by E. coli is 10.80 times more than that produced in darkness. The findings strongly indicate that industrial microbial waste, whether eukaryote or prokaryote, can be employed for the economical production of nano-Se or other nanoparticles.
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INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
·
2026
·
IF 8.7
Lin-hui Li,Meng-Sheng Wei,Jia-Ren Cao,Yu-Qiu Li,Wei Tao,Wen-Ning Feng,Bo Liu,Ling-Qia Su
Polyethylene terephthalate (PET) pollution poses a global environmental challenge, and enzymatic degradation represents a promising eco-friendly solution. The BhrPETase mutant Bhr6M demonstrated high degradation efficiency, showing considerable potential for PET biodegradation. To facilitate its practical application, we scaled up recombinant Bhr6M production in Escherichia coli via a 3-L bioreactor and identified optimal induction conditions of 30 °C and lactose feeding at 0.3 g·L −1 ·h −1 . However, the enzyme solution exhibited limited stability, with a half-life ( t 1/2 ) of approximately four days at 30 °C and four hours at 70 °C, hindering its broader application. To address this issue, we rationally designed via AlphaFold3, PROSS and combinatorial mutagenesis, obtaining the double mutant Bhr-S184H/M57I with synergistic performance improvements. This mutant achieved a yield of 3.17 g·L −1 in approximately 31 h of fermentation, representing the highest productivity reported for PET hydrolases. Its stability was enhanced with a 66-fold longer t 1/2 at 70 °C than Bhr6M, and over 90% residual activity after 30 days at 30 °C. In addition, the mutant degraded 97% of amorphous PET at 70 °C within 48 h, a process that required 96 h for Bhr6M at its optimal temperature of 60 °C. After that, we further validated its practical utility by achieving over 95% degradation of pretreated PET bottles. Molecular dynamics (MD) simulations revealed the enhanced stability originates from new hydrogen-bonding networks and reduced conformational flexibility in key regions. This study provides a scalable strategy to simultaneously improve enzyme yield and stability, laying a foundation for industrial PET biodegradation.
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Microorganisms
·
2026
·Vol. 14
·期 5
·1088
·
IF 4.7
Yuyang Liu,Yaxin Wen,Ruizheng Hu,Ruyue Han,Dong Liu,Hailing Zhang
Ergothioneine (EGT), a naturally occurring amino acid derivative with potent antioxidant and cytoprotective properties, is widely applied in the food, cosmetic, and medical industries. Traditional production methods are limited by high costs, low efficiency, and environmental concerns, so microbial fermentation serves as a sustainable alternative for EGT production. In this study,Escherichia coliBL21 (DE3) was employed as the chassis strain. First, a basic EGT-producing engineered strain was constructed by heterologously expressing theegtBgene fromMethylobacterium pseudosasicolaalong with theegtDandegtEgenes fromMycobacterium smegmatis. This initial strain achieved a yield of 84.84 ± 1.64 mg/L of EGT in shake-flask cultures. To enhance production, solubility-enhancing tags were introduced to improve the soluble expression of the key enzymes, and metabolic pathways were rationally engineered to strengthen the supply of essential precursor amino acids. These modifications led to the development of a high-yield EGT strain. After optimizing the fermentation process, the best results were achieved using a medium with glycerol as the carbon source, 0.5 g/L of histidine, 1.5 g/L of methionine, and 1.0 g/L of cysteine, along with induction at 25 °C using 0.2 mM IPTG for 120 h. Under these conditions, the final EGT yield reached 385.70 ± 4.86 mg/L. The engineered strain for EGT synthesis and optimized fermentation strategy developed in this study offer a useful basis for further process development.
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JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
·
2026
·
IF 6.7
Cheng Chen,Yamin Huang,Juan Li,Meijuan Xu,Hui Li,Guoqiang Xu,Jinsong Shi,Zhenghong Xu,Xiaomei Zhang
l-Tryptophan, an essential amino acid, serves as a critical precursor for the biosynthesis of numerous bioactive compounds. In this study, we engineered tryptophan synthase (TrpS) to develop an efficient cell factory for l-tryptophan production. Initially, EcTrpS from Escherichia coli K-12 was expressed in E. coli BL21 (DE3) to construct strain BS, which produced 836.43 mg/L of l-tryptophan. To alleviate substrate inhibition, we constructed a mutant library of 104–105 variants through two rounds of error-prone PCR and utilized a biosensor employing a green fluorescent protein to report l-serine consumption for screening TrpS mutants. Mutant C10 showed a 375% increase in enzyme activity compared with strain BS. Computational analysis was used to explore an improved catalytic mechanism. Finally, through systematic process optimization and catalytic strategy, strain C10 produced 94.2 g/L of l-tryptophan, the highest titer via whole-cell transformation. This study provides new perspectives and strategies for TrpS engineering and the industrial production of l-tryptophan.
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ACS Synthetic Biology
·
2026
·
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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npj Clean Water
·
2023
·Vol. 6
·期 1
·1-11
·
IF 11.4
Li Chao,Li Jiale,Huang Niu,Kong Xin Ying,Xiao Qingyi,Huang Yingping,Wong Po Keung,Ye Liqun
Advanced oxidation technology based on peroxonosulfate (PMS) has attracted extensive attention in water treatment research due to its fast reaction speed and wide pH range adaptability. Cobalt-based catalysts are considered to be one of the most effective reagents for PMS activation in various PMS activation methods. However, Co-ion leaching and difficulty in recovery have greatly hindered its practical applications. Herein, we developed a robust membrane constructed by nitrogen-doped carbon nanotubes embedded with cobalt nanoparticles (Co-NC) to concurrently address the Co-ion leaching and recovery issues. Based on our customization, continuous water flow reactor, the Co-NC membrane exhibited excellent catalytic activity and stability, in which it demonstrated a remarkable sterilization efficiency of 99.9999% against E. coli , and it retained a superior stability of 96.29% after 40 repeated cycles. Fewer attempts to put such efficient heterogeneous advanced oxidation processes (AOPs) into practical application, to mimic real-life applications, the performance of the Co-NC/PMS system was extended to the water taken from Qiuxi River. Remarkably, there is no deterioration in performance over 12 h of continuous real sewage processing. Mechanistic studies revealed that abundant high-valence metals (Co IV =O) were generated in the system, which can attack and penetrate into the cell membrane to destroy its intracellular defense system. This work provides useful insights into designing robust membranes with superior efficiency and stability for PMS-based advanced oxidation technology.
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BIOPROCESS AND BIOSYSTEMS ENGINEERING
·
2022
·Vol. 45
·期 3
·563-571
·
IF 3.434
Mei Jianfeng,Wu Xia,Zheng Sujing,Yi Yu,Wang Xudong,Ying Guoqing
Bilirubin, a natural intermediate in heme degradation, is a valuable Chinese medicine used in more than 50 traditional Chinese medicine (TCM) preparations. At present, bilirubin is mainly produced by extraction from pig bile, but a shortage of the raw material has increased the price, to about US$10,000/kg in the Chinese market. Biliverdin, the precursor of bilirubin, is more abundant and less expensive than bilirubin, but it is not used in TCM. Thus, the biotransformation of biliverdin by biliverdin reductase (BvdR) may be a practical way to produce bilirubin. In this study, the codon-optimized gene of biliverdin reductase ( m bvdR ) from the cyanobacterium Synechocystis was cloned into Escherichia coli BL21(DE3), and the conditions for BL21-mBvdR expressing BvdR were optimized. Resting BL21-mBvdR cells were employed as biocatalysts to biotransform biliverdin to bilirubin. At a concentration of biliverdin substrate of 450 mg/L in the reaction mixture, the bilirubin content in dry cells reached 20.8 ± 0.8 mg/g, with a conversion yield of 72.3%. Therefore, recombinant E. coli expressing BvdR can be applied to biotransform biliverdin to bilirubin, providing a potential alternative process for bilirubin production. Graphical abstract
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Journal of Water Process Engineering
·
2023
·
IF 7
Bing Li,Chiqian Zhang,Qian Zuo,Huaitian Bu,Jinxiu Li,Ping Li,Jinhua Wu
In this work, α-FeOOH modified activated carbon (FeOOH@AC) was prepared through wetness-impregnation and used as a persulfate (S 2 O 8 2− , PS) activator to inactivate Escherichia coli in water. Surficial analysis displayed that micron-sized (0.5 to 1 μm) goethite was evenly distributed on AC. The structural ferric ions in the FeOOH matrix were partially reduced to ferrous ions by the hydroxyl of AC. The surface-bonded Fe 2+ facilitated PS to produce hydroxyl and sulfate radicals (·OH and SO 4 · − ). These radicals effectively inactivated Escherichia coli by destructing bacterial membrane. Therefore, the FeOOH@AC-PS system obtained a higher inactivation rate of 6.0 log as compared to 3.1 log and 0.4 log in the AC-PS and FeOOH-PS systems, respectively. After sterilization, the Escherichia coli did not regrow in the FeOOH@AC-PS system within 48 h, indicating its persistent growth inhibition by radicals. The FeOOH@AC-PS system also showed high inactivation efficiency over a pH range of 3 to 11 and excellent stability in cyclic test. Spent FeOOH@AC was amenable for effective regeneration by eliminating surficial iron oxides with hydrochloric acid and reloading FeOOH through wetness-impregnation with a regeneration rate up to 96.7 %. These results demonstrated that FeOOH@AC can effectively and stably activate PS for inactivating bacteria in water.
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BIORESOURCE TECHNOLOGY
·
2023
·
IF 11.4
Zixin Teng,Xuewei Pan,Yunran Liu,Jiajia You,Hengwei Zhang,Zhenqiang Zhao,Zhina Qiao,Zhiming Rao
L-serine is a high-value amino acid widely used in the food, medicine, and cosmetic industries. However, the low yield of L-serine has limited its industrial production. In this study, a cellular factory for efficient synthesis of L-serine was obtained by engineering the serine hydroxymethyltransferases (SHMT). Firstly, after screening the SHMT from Alcanivorax dieselolei by genome mining, a mutant Ad SHMT E266M with high thermal stability was identified through rational design. Subsequently, an iterative saturating mutant library was constructed by using coevolutionary analysis, and a mutant Ad SHMT E160L/E193Q with enzyme activity 1.35 times higher than Ad SHMT was identified. Additionally, the target protein Ad SHMT E160L/E193Q/E266M was efficiently overexpressed by improving its mRNA stability. Finally, combining the substrate addition strategy and system optimization, the optimized strain BL21/pET28a- Ad SHMT E160L/E193Q/E266M -5′UTR-REP3 S16 produced 106.06 g/L L-serine, which is the highest production to date. This study provides new ideas and insights for the engineering design of SHMT and the industrial production of L-serine.
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ECOTOXICOLOGY AND ENVIRONMENTAL SAFETY
·
2020
·
IF 4.872
Qiao Zhong,Ying Deng,Huaming Qin,Huase Ou,Yanfen Qu,Jinshao Ye
Although the toxicity of triclocarban at molecular level has been investigated, the metabolic networks involved in regulating the stress processes are not clear. Whether the cells would maintain specific phenotypic characteristics after triclocarban stress is also needed to be clarified. In this study, Escherichia coli was selected as a model to elucidate the cellular metabolism response associated with triclocarban stress and the recovery metabolic network of the triclocarban-treated cells using the proteomics and metabolomics approaches. Results showed that triclocarban caused systematic metabolic remodeling. The adaptive pathways, glyoxylate shunt and acetate-switch were activated. These arrangements allowed cells to use more acetyl-CoA and to reduce carbon atom loss. The upregulation of NH 3 -dependent NAD + synthetase complemented the NAD + consumption by catabolism, maintaining the redox balance. The synthesis of 1-deoxy-D-xylulose-5-phosphate was suppressed, which would affect the accumulation of end products of its downstream pathway of isoprenoid synthesis. After recovery culture for 12 h, the state of cells returned to stability and the main impacts on metabolic network triggered by triclocarban have disappeared. However, drug resistance caused by long-term exposure to environmentally relevant concentration of triclocarban is still worthy of attention. The present study revealed the molecular events under triclocarban stress and clarified how triclocarban influence the metabolic networks.
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