Biotechnology for Biofuels and Bioproducts
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2022
·Vol. 15
·期 1
·1-12
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IF 0
Hu Yueli,Zhang Chen,Zou Lihua,Zheng Zhaojuan,Ouyang Jia
Background The conversion of lignin-derived aromatic monomers into valuable chemicals has promising potential to improve the economic competitiveness of biomass biorefineries. Pinosylvin is an attractive pharmaceutical with multiple promising biological activities. Results Herein, Escherichia coli was engineered to convert the lignin-derived standard model monomer cinnamic acid into pinosylvin by introducing two novel enzymes from the wood plant: stilbene synthase from Pinus pinea (PpSTS) and 4-Coumarate-CoA ligase from Populus trichocarpa (Ptr4CL4) . The expression of Ptr4CL4 drastically improved the production of pinosylvin (42.5 ± 1.1 mg/L), achieving values 15.7-fold higher than that of Ptr4CL5 (another 4-Coumarate-CoA ligase from Populus trichocarpa ) in the absence of cerulenin. By adjusting the expression strategy, the optimized engineered strain produced pinosylvin at 153.7 ± 2.2 mg/L with an extremely high yield of 1.20 ± 0.02 mg/mg cinnamic acid in the presence of cerulenin, which is 83.9% ± 1.17 of the theoretical yield. This is the highest reported pinosylvin yield directly from cinnamic acid to date. Conclusion Our work highlights the feasibility of microbial production of pinosylvin from cinnamic acid and paves the way for converting lignin-related aromatics to valuable chemicals.
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Gut Microbes
·
2023
·
IF 12.2
Jie Hu,Yifan Wu,Xingjian Zhou,Luyuan Kang,Shiyi Zhang,Yisi Liu,Yu Pi,Xilong Li,Junjun Wang,Dandan Han
The pathogenicity of Escherichia coli (E. coli) O157:H7 is predominantly associated with Shiga toxin 2 (Stx2) that poses a huge threat to human and animal intestinal health. Production of Stx2 requires expression of stx2 gene, which is located in the genome of lambdoid Stx2 prophage. Growing evidence has implicated that many commonly consumed foods participate in the regulation of prophage induction. In this study, we aimed to explore whether specific dietary functional sugars could inhibit Stx2 prophage induction in E. coli O157:H7, thereby preventing Stx2 production and promoting intestinal health. We demonstrated that Stx2 prophage induction in E. coli O157:H7 was strongly inhibited by L-arabinose both in vitro and in a mouse model. Mechanistically, L-arabinose at doses of 9, 12, or 15 mM diminished RecA protein levels, a master mediator of the SOS response, contributing to reduced Stx2-converting phage induction. L-Arabinose inhibited quorum sensing and oxidative stress response, which are known as positive regulators of the SOS response and subsequent Stx2 phage production. Furthermore, L-arabinose impaired E. coli O157:H7 arginine transport and metabolism that were involved in producing Stx2 phage. Collectively, our results suggest that L-arabinose may be exploited as a novel Stx2 prophage induction inhibitor against E. coli O157:H7 infection.
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APPLIED BIOCHEMISTRY AND BIOTECHNOLOGY
·
2022
·Vol. 194
·期 12
·5977-5991
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IF 3.094
Fan Xianyu,Qi Zhipeng,Zhang Xiaomeng,Pei Jianjun,Zhao Linguo
Pseudoionone is a valuable intermediate for the flavor industry. However, few attempts have been made to synthesize pseudoionone using biotechnology. In this work, an Escherichia coli strain harboring both the isopentenol utilization pathway (IUP) and mevalonate (MVA) pathway was engineered for the production of the pseudoionone precursor lycopene, which increased the titer of lycopene to approximately 20-fold the yield obtained by the original MEP pathway. Subsequently, the crucial limiting step of carotenoid cleavage dioxygenases (CCDs) was evaluated and optimized. The most effective upstream module for pseudoionone synthesis in E. coli was determined to be MnCCD1 from Morus notabilis without a GST-tag and under the control of the tac promoter. Finally, the highest pseudoionone production achieved 20.61 mg/L under the optimum fermentation conditions in a shake flask. This study provided an efficient approach for pseudoionone production and advanced our understanding of the characteristics of the CCD family for the biosynthesis of aroma compounds.
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Fermentation-Basel
·
2023
·Vol. 9
·期 8
·764
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IF 3.7
Xinchao Yang,Linlin Niu,Chunjiang Ye,Yuanxiu Wang,Yuehui Liu,Fang Wang,Naxin Sun
D-glucaric acid is an important bio-based building block of polymers and is a high value-added chemical that can be used in a variety of applications. In the present study, theUdhtarget genes fromPseudomonas putidaandPseudomonas syringaewere used together to construct the expression vector pETDuet-2 × Udh. The transformants of BL21 (DE3) with vector pETDuet-2 × Udh were applied to produce glucaric acid from glucuronic acid. After optimizing the induction conditions, the highest Udh expression was achieved when 0.4 mmol·L−1isopropyl-β-d–thiogalactoside (IPTG) was added to the cell cultures at an OD600value of 0.6 followed by culturing at 26 °C for 6 h. The production of glucaric acid substantially reached 5.24 ± 0.015 g·L−1in fed-batch cultures in a 30 L tank. In the present study, a new system for glucaric acid production was established, which was more economic and friendly to the environment.Keywords:urinate dehydrogenases;pETDuet-2 × Udh;D-glucaric acid;glucuronic acid;SDS-PAGE
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Microbial Cell Factories
·
2022
·Vol. 21
·期 1
·1-14
·
IF 6.352
Liu Yan,Wang Xiaoliang,Nong Sujin,Bai Zehui,Han Nanyu,Wu Qian,Huang Zunxi,Ding Junmei
Background Carbamate pesticides have been widely used in agricultural and forestry pest control. The large-scale use of carbamates has caused severe toxicity in various systems because of their toxic environmental residues. Carbaryl is a representative carbamate pesticide and hydrolase/carboxylesterase is the initial and critical enzyme for its degradation. Whole-cell biocatalysts have become a powerful tool for environmental bioremediation. Here, a whole cell biocatalyst was constructed by displaying a novel carboxylesterase/hydrolase on the surface of Escherichia coli cells for carbaryl bioremediation.Results The carCby gene, encoding a protein with carbaryl hydrolysis activity was cloned and characterized. Subsequently, CarCby was displayed on the outer membrane of E. coli BL21(DE3) cells using the N-terminus of ice nucleation protein as an anchor. The surface localization of CarCby was confirmed by SDS–PAGE and fluorescence microscopy. The optimal temperature and pH of the engineered E. coli cells were 30 °C and 7.5, respectively, using pNPC4 as a substrate. The whole cell biocatalyst exhibited better stability and maintained approximately 8-fold higher specific enzymatic activity than purified CarCby when incubated at 30 °C for 120 h. In addition, ~ 100% and 50% of the original activity was retained when incubated with the whole cell biocatalyst at 4 ℃ and 30 °C for 35 days, respectively. However, the purified CarCby lost almost 100% of its activity when incubated at 30 °C for 134 h or 37 °C for 96 h, respectively. Finally, approximately 30 mg/L of carbaryl was hydrolyzed by 200 U of the engineered E. coli cells in 12 h.Conclusion sHere, a carbaryl hydrolase-containing surface-displayed system was first constructed, and the whole cell biocatalyst displayed better stability and maintained its catalytic activity. This surface-displayed strategy provides a new solution for the cost-efficient bioremediation of carbaryl and could also have the potential to be used to treat other carbamates in environmental bioremediation.
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SENSORS AND ACTUATORS B-CHEMICAL
·
2021
·
IF 7.46
Yadan Guo,Yi Luo,Minggen Tang,Meng Zhang,Mao-sen Yuan,Shuwei Chen,Qin Tu,Jinyi Wang
A promising gold nanorod sensor that demonstrated comparable antibacterial ability, selectively recognized Escherichia coli ( E. coli ), and used selenol as the recognition group was prepared and applied for foodstuff E.coli detection. We used a sulfhydryl compound linked with fluorescent dye by sulfamide bonds ( SH-NB ) to modify gold nanorods ( GNRs ) and fabricate GNRs-SH-NB that can selectively and sensitively detect selenol. Selenol group broke sulfamide bonds, causing the fluorescent dye to fall off from GNRs-SH-NB and exhibit red fluorescence, demonstrating that it can selectively identify E. coli with the unique selenoenzymes or selenoproteins in the bacterium. After the decomposition of GNRs-SH-NB , the obtained GNRs modified by N-heterocyclic sulfhydryl compounds and positively charged fluorescent dyes showed synergistic antibacterial properties. Furthermore, the photothermal effect of GNRs increased the temperature beyond the optimum temperature of E. coli and caused its maladjustment, leading to the death of the bacterium. Thus, GNRs-SH-NB exerted a considerable antibacterial effect against E. coli and accurately detected E. coli in milk, meat, and eggshell. With the low limit of detection, this novel antibacterial sensor GNRs-SH-NB provide a promising method to identify the E. coli and further utilized in food safety field initially.
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NEW JOURNAL OF CHEMISTRY
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2020
·Vol. 44
·期 41
·17671-17678
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IF 3.288
Saira Shams,Waqas Ahmad,Amjad Hussain Memon,Sumaira Shams,Yun Wei,Qipeng Yuan,Hao Liang
The objective of this study was to design a more effective antibacterial agent to overcome the problem of fast-growing bacterial resistance. The results showed that a Cu/H3BTC metal–organic framework (MOF) exhibited a notable antibacterial performance towards Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli), and could significantly disrupt the cell membrane, resulting in the discharge of cell constituents. In addition, morphological changes were elucidated using scanning electron microscopy (SEM). SEM photographs demonstrated noticeable and apparent changes in the morphology of bacterial cells, signifying cell damage that was further confirmed via confocal laser scanning microscopy (CLSM). Furthermore, the result of agarose gel electrophoresis (AGE) showed that Cu/H3BTC MOF could enter the bacterial cells by impairing cell membranes and inhibiting the synthesis of DNA. These findings indicated that the Cu/H3BTC MOF is a potent antimicrobial material that can be applied to hinder the growth of pathogenic microbes.
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ENVIRONMENTAL POLLUTION
·
2020
·
IF 6.793
Dali Wang,Qing Ning,Jiayu Dong,Bryan W. Brooks,Jing You
Antibiotics in the environment usually co-exist with their transformation products with retained toxicity, raising concerns about environmental risks of their combined exposure. Herein, we reported a novel predictive approach for evaluating the individual and combined toxicity for photodegradation products of fluoroquinolone antibiotics (FQs). Quantitative structure-activity relationship (QSAR) models with promising predictive performance were constructed and validated using experimental data obtained with 13 FQs and 78 mixtures towards E. coli . A structural descriptor reflecting the interaction among FQ molecules and the target protein was employed in the QSAR models, which was obtained through molecular docking and thus provided a rational mechanistic explanation for these models. The predicted results indicated that the degradation products displayed varying degrees of changes compared to the parent FQs, while the combined toxicity of FQs and their degradation products was mostly additive. Furthermore, following UV irradiation the degradation products displayed elevated capacity of inducing resistance mutations in E. coli , though their overall toxicity was reduced. This result highlights the implications of antibiotic degradation products on resistance development in bacteria and stresses the importance of considering such impacts during environmental risk assessments of antibiotics.
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PROCESS BIOCHEMISTRY
·
2021
·
IF 3.757
Yangyang Cai,Shuqi Xing,Qifeng Zhang,Ruonan Zhu,Kai Cheng,Cuiqin Li,Xuefeng Zeng,Laping He
The extracellular Aspergillus niger GZUF36 lipase (EXANL1) has broad applicability such as synthesis of functional lipids. The wild-type strain is unstable, and the culture time is long. Studies on the selectivity of substrates for EXANL1 are not available. Therefore, this study aimed to express, purify, characterize and analyze the substrate specificity of EAXNL1. The results showed that EXANL1 was stable in the presence of 40 % ( v/v ) dimethyl sulfoxide, glycerol, toluene, and n-hexane. The relative activity was 104 %, 88 %, 81 %, and 76 %. The residual enzyme activity of EXANL1 was more than 85 % in surfactants with a low concentration. EXANL1 exhibited a high conversion rate (79 %) for p -nitrophenyl palmitate. The molecular docking of p -nitrophenyl palmitate into the active site of EXANL1 indicated that the preference for this fatty acid chain length might be due to the formation of hydrogen bonds. The distance between the carbonyl carbon atom of p -nitrophenyl palmitate and the nucleophilic oxygen atom of serine 162 was 2.91 Å. Both experiments revealed that EXANL1 exhibited higher specificity for p -nitrophenyl palmitate. These results indicated that EXANL1 could be a promising tool in organic synthesis and detergent industry. The molecular docking provided a basis for the explanation on the substrate selectivity of EXANL1.
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JOURNAL OF CHEMICAL TECHNOLOGY AND BIOTECHNOLOGY
·
2018
·Vol. 94
·期 1
·159-166
·
IF 2.587
Jun Li,Miaoliang Fan,Rongjin Zhang,Zheyu Tian,Jing Zeng,Yinan Song,Wenting Du
Background Ionic liquids (ILs) have advantages in terms of flexible structure and composition for the whole cell mediated biocatalytic reaction. However, the lack of design strategy for ILs has hampered the development of biocatalysis in non-aqueous medium. In the present study, to establish an efficient reaction system to obtain vital chiral compounds, a novel IL was designed by combining the superior characteristics of quaternary ammonium- and dialkyl-based cationic surfactant IL to enhance the recombinant Escherichia coli cells-catalyzed bioreduction efficiency. Results [HOEtN1,1,1][BF 4 ] was synthesized and showed excellent characteristics of low toxicity to the biocatalyst, reduced toxicity of substrate to the microbial cells and moderate increase of cell membrane permeability. In the [HOEtN1,1,1][BF 4 ]- containing reaction system, some valuable factors involved in the biocatalytic reduction of ethyl 4-chloro-3-oxobutanoate to ethyl ( S )-4-chloro-3-hydroxybutanoate were found to be: 2.5% (w/v) [HOEtN1,1,1][BF 4 ], 20% (v/v) isopropanol, 3 M COBE, pH 7.0, 30 °C, reacted for 16 h. A yield of 96.7% was obtained under the optimum conditions with above 99% enantiomeric excess value, and the proposed bioprocess was also successfully scaled up to a 5 L fermentor with a yield of 90.7%. Conclusion The developed IL [HOEtN1,1,1][BF 4 ] is a promising solvent for enhancing the efficiency of model bioreduction. It was also evaluated for broadening its application to the biosynthesis of ( R )-[3,5-bis(trifluoromethyl)phenyl] ethanol catalyzed by recombinant Escherichia coli , with remarkable success. © 2018 Society of Chemical Industry
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NANOTECHNOLOGY
·
2018
·Vol. 30
·期 2
·025501
·
IF 3.404
Danqing Liu,Tianchan Li,Weicheng Huang,Zhuang Ma,Wei Zhang,Rui Zhang,Hong Yan,Bin Yang,Shaoqin Liu
Foodborne pathogens are perpetual threats to human and animal health. Detection of pathogens requires accurate, sensitive, rapid and point-of-care diagnostic assays. In this study, we described a simple and sensitive electrochemiluminescent (ECL) assay to detect the deadly bacteria Escherichia coli O157:H7 by {\rm{Ru}}{{({\rm{bpy}})}_{3}}^{2+}-coated ZnO nanorods arrays (NAs). The {\rm{Ru}}{{({\rm{bpy}})}_{3}}^{2+}-coated ZnO NAs were fabricated by immobilizing {\rm{Ru}}{{({\rm{bpy}})}_{3}}^{2+} on ZnO NAs with a large specific surface area and good conductivity. An {\rm{Ru}}{{({\rm{bpy}})}_{3}}^{2+}-2-(dibutylamino)-ethanol (DBAE) system coated on ZnO NAs exhibits high ECL intensity, rapid response and good stability. This system was further developed as an ECL immunosensor used in the detection of E. coli O157:H7. The proposed ECL immunosensor exhibits a broad detection range within the scope of 200–100 000 CFU ml−1 and quite a low detection limit of 143 CFU ml−1. The high specificity, remarkable reproducibility and good stability offer a sensitive, selective, and convenient pathway for detecting E. coli O157:H7 in the field of food safety and clinical diagnosis.
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SEPARATION AND PURIFICATION TECHNOLOGY
·
2018
·
IF 3.927
Haoran Dong,Yujun Cheng,Yue Lu,Kunjie Hou,Lihua Zhang,Long Li,Bin Wang,Yaoyao Wang,Qin Ning,Guangming Zeng
This study examined the toxicities of Fe/Ni and starch-stabilized Fe/Ni (S-Fe/Ni) nanoparticles (NPs) toward Escherichia coli . The inactivation of E. coli by Fe/Ni NPs was concentration- and time-dependent. Fe/Ni NPs exhibited a stronger toxicity than the bare Fe 0 NPs (i.e., Ni = 0 wt%), revealing that the loading of Ni increased the toxicity of Fe 0 NPs. However, there was no positive correlation between Ni content and the toxicity of Fe/Ni NPs when Ni content increased from 1 wt% to 5 wt%. The starch coating could alleviate the toxicity of Fe/Ni NPs, depending on starch concentration. Transmission Electron Microscopy analysis demonstrates that membrane disruption and cellular internalization of NPs occurred for E. coli after exposure to the Fe/Ni NPs. The NPs might have reacted with the carboxyl, ester and amine groups on the cell surface, as indicated by the Fourier Transform Infrared Spectroscopy analysis. However, the S-Fe/Ni NPs did not cause obvious membrane disruption of the cells due to the effect of starch coating, which inhibited the close contact of NPs with cells by providing electrostatic repulsion and steric hindrance. Besides, the addition of N-Acetyl-L-cysteine, as a potent scavenger for reactive oxygen species (ROS), remarkably reduced the toxicities of both Fe/Ni and S-Fe/Ni NPs, which indicated that the ROS-induced oxidative stress should be the primary mechanism for the inactivation of E. coli . The aging experiments evidenced that both Fe/Ni and S-Fe/Ni NPs would lose their toxic potential over time, posing less threat to the environment in the long term.
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