Frontiers in Microbiology
·
2021
·
IF 5.64
Zou Lei,Li Sha,Li Nan,Ruan Shi-Long,Chen Jing,Wu Jing,Yan Dazhong,Chao Hong-Jun
Escherichia coli has been developed as the most common host for recombinant protein expression. Unfortunately, there are still some proteins that are resistant to high levels of heterologous soluble expression in E. coli. Protein and peptide fusion tags are one of the most important methods for increasing target protein expression and seem to influence the expression efficiency and solubility as well. In this study, we identify a short 15-residue enhancing solubility peptide, the PCDS (protocatechuate 3,4-dioxygenase solubility) tag, which enhances heterologous protein expression in E. coli. This PCDS tag is a 45-bp long sequence encoding a peptide tag involved in the soluble expression of protocatechuate 3,4-dioxygenase, encoded by the pcaHG98 genes of Pseudomonas putida NCIMB 9866. The 45-bp sequence was also beneficial for pcaHG98 gene amplification. This tag was shown to be necessary for the heterologous soluble expression of PcaHG98 in E. coli. Purified His6-PcaHG98e04-PCDS exhibited an activity of 205.63±14.23U/mg against protocatechuate as a substrate, and this activity was not affected by a PCDS tag. This PCDS tag has been fused to the mammalian yellow fluorescent protein (YFP) to construct YFP-PCDS without its termination codons and YFPt-PCDS with. The total protein expressions of YFP-PCDS and YFPt-PCDS were significantly amplified up to 1.6-fold and 2-fold, respectively, compared to YFP alone. Accordingly, His6-YFP-PCDS and His6-YFPt-PCDS had 1.6-fold and 3-fold higher soluble protein yields, respectively, than His6-YFP expressed under the same conditions. His6-YFP, His6-YFP-PCDS, and His6-YFPt-PCDS also showed consistent fluorescence emission spectra, with a peak at 530nm over a scanning range from 400 to 700nm. These results indicated that the use of the PCDS tag is an effective way to improve heterologous protein expression in E. coli.
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CyTA-Journal of Food
·
2024
·
IF 2
Liyuan Niu,Dianhe Zhang,Bohua Wang,Qisen Xiang,Yanhong Bai
This study investigated the inactivation mechanism and kinetics of dielectric barrier discharges (DBD) plasma against Escherichia coli O157:H7. DBD plasma exhibited a remarkable bactericidal effect, and the inactivation kinetics was well described with the Weibull model. Using the ambient air as the working gas, surviving E. coli O157:H7 in phosphate-buffered saline achieved an undetectable level (<10 CFU/mL) after treatment at 18 W for 60 s. DBD plasma altered cell morphology, destructed both the outer and inner membranes, and increased intracellular reactive oxygen species levels. Moreover, DBD plasma significantly decreased the pH of the suspended medium, and increased its oxidation reduction potential and active species levels, which could be mainly responsible for microbial inactivation. The presence of glutathione or N-acetyl-L-cysteine weakened the antibacterial activity of DBD plasma due to their reactive oxygen and nitrogen species (RONS) scavenging capacity, demonstrating that RONS generated by DBD plasma played important roles in decontamination.
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Bioresource Technology Reports
·
2021
·
IF 0
Ya Liu,Rui Pei,Zhongliang Huang,Jingran Xiao,Ayan Yao,Kaiji Xu,Yixin Li,Shafqat Ullah,Zongzhong Yu,Yuanpeng Wang,Shu-Feng Zhou,Guowu Zhan
Herein, we propose a facile synthetic strategy for biogenic photocatalyst by sequentially immobilizing CdS and Pt nanoparticles (NPs) on engineered recombinant Escherichia coli ( e-E. coli ) strains. In particular, we transferred the cysteine desulfurase gene to the heterologous host to overexpress cysteine desulfurase and the resulting e-E. coli strain could serve as a biofactory for the extracellular growth of CdS NPs involving catalyzing L-cysteine (Cys) to form a reactive sulfur source, followed by the precipitation of CdS via the reaction between Cd 2+ ions and the locally concentrated active sulfur source around the strain. The subsequent modification of e-E. coli /CdS composites with noble metal Pt NPs further enhanced the catalytic degradation performance towards organic dyes, due to suppressing the recombination of the photogenerated electron-hole pairs. Accordingly, the ternary e-E. coli /CdS/Pt composites emerged as robust photocatalysts for pollutant degradation in waste-water treatment, which is promising for environmental remediation in ambient conditions.
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DOI
SENSORS
·
2025
·Vol. 25
·期 8
·2457
·
IF 3.5
Deying Wang,Yan Chen,Qi Zhang,Junfei Chen,Changhao Li,Yunjing Luo,Yong Jin,Xiaohua Qi
HighlightsWhat are the main findings?Developed a novel WS2-Au nanocomposite coupled with 5,5′-dithiobis-(2-nitrobenzoic acid) (DTNB) and antibodies as sensitive surface-enhanced Raman scattering (SERS) nanotags forE. coliO157:H7 detection.Demonstrated a broad linear detection range (8 × 102–8 × 107CFU/mL), a low detection limit (175 CFU/mL), high specificity and good accuracy in real food samples (milk, pork).What are the implications of the main findings?A rapid assay for qualitative and quantitative detection ofE. coliO157:H7, overcoming limitations of traditional ICA methods.A cost-effective, user-friendly surface-enhanced Raman scattering (SERS)-based immunochromatographic assay (SERS-ICA) platform with great potential for point-of-care testing and detection of other foodborne pathogens.E. coliO157:H7 contamination in food and the environment poses a serious threat to human health. Rapid and sensitive identification of foodborne pathogens remains challenging. Here, we prepared tungsten disulfide (WS2)–Au nanocomposites coupled with the Raman signal molecule 5,5′-dithio-bis-(2-nitrobenzoic acid) (DTNB) and antibodies to replace the conventional colloidal gold nanoparticles and applied SERS-active nanotags in the SERS-ICA method for highly sensitive detection ofE. coliO157:H7. The large surface area and numerous effective SERS hotspots of WS2-Au nanotags provide superior SERS signals. Under optimized conditions, this ICA achieves the quantitative detection ofE. coliO157:H7 in a broad linear range of 8 × 102–8 × 107CFU/mL and at a low detection limit of 175 CFU/mL. In addition, the test strip indicates high specificity forE. coliO157:H7 identification, favorable reproducibility, and shows good accuracy in the detection of actual food samples, such as milk and pork. The proposed assay can be used for rapid qualitative and quantitative detection ofE. coliO157:H7 and has great potential for field application.
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mSphere
·
2024
·
IF 3.7
Xiao-die Cui,Xiao-kang Liu,Xiao-yuan Ma,Shuai-hua Li,Jun-kai Zhang,Rong-jia Han,Kai-fang Yi,Jian-hua Liu,Yu-shan Pan,Dan-dan He,Gong-zheng Hu,Ya-jun Zhai
The appearance and prevalence of multidrug-resistance (MDR) Gram-negative bacteria (GNB) have limited our antibiotic capacity to control bacterial infections. The clinical efficacy of colistin (COL), considered as the “last resort” for treating GNB infections, has been severely hindered by its increased use as well as the emergence and prevalence of mobile colistin resistance (MCR)-mediated acquired drug resistance. Identifying promising compounds to restore antibiotic activity is becoming an effective strategy to alleviate the crisis of increasing MDR. We first demonstrated that the combination of berberine (BBR) and EDTA substantially restored COL sensitivity against COL-resistant Salmonella and Escherichia coli. Molecular docking indicated that BBR can interact with MCR-1 and the efflux pump system AcrAB-TolC, and BBR combined with EDTA downregulated the expression level of mcr-1 and tolC. Mechanically, BBR combined with EDTA could increase bacterial membrane damage, inhibit the function of multidrug efflux pump, and promote oxidative damage, thereby boosting the action of COL. In addition, transcriptome analysis found that the combination of BBR and EDTA can accelerate the tricarboxylic acid cycle, inhibit cationic antimicrobial peptide (CAMP) resistance, and attenuate Salmonella virulence. Notably, the combination of BBR and EDTA with COL significantly reduced the bacterial load in the liver and spleen of a mice model infected with Salmonella. Our findings revealed that BBR and EDTA can be used as adjuvants collectively with COL to synergistically reverse the COL resistance of bacteria.
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BIOCHEMICAL ENGINEERING JOURNAL
·
2025
·
IF 3.8
Xue-Chun Li,Hong-Jie Xie,Zheng-Xiong Zhou,Bin Sun,Jun-Jie Lin,Xiao-Dan Li,Ren Wang
1-aminocyclopropane-1-carboxylic acid (ACC), a crucial intermediate in the ethylene biosynthetic pathway, plays a decisive role in regulating plant developmental processes and stress responses. In heterologous biosynthesis systems, ACC biosynthesis is inhibited due to its antibacterial activity. Enhancing ACC export confers advantages to ACC-producing biotechnological platforms and microbial cell factories. While bacterial transporters for canonical amino acids in Corynebacterium glutamicum and Escherichia coli have been extensively studied, their roles in ACC transport remain unexplored. To identify the exporters mediating ACC transport in E. coli , relevant experiments were performed in this study. The results showed that the Leucine-exporter LeuE acts as a high-efficiency ACC exporter in E. coli . Subsequently, structural modeling coupled with site-directed mutagenesis revealed that L160 is a critical residue in determining the ACC transport specificity of LeuE. The L160Y mutation enhanced E. coli tolerance to ACC, elevating the threshold from 50 mM to 100 mM. Structural modeling predicted that the L160Y mutation-induced compaction reduced the transporter channel diameter by 28% (from 9.551 Å to 6.876 Å), a change that may enhance the binding affinity of ACC. Notably, a significant reduction in intracellular ACC levels was observed in both the LeuE-overexpressing and LeuE (L160Y)-overexpressing strains. These findings enable large-scale biosynthesis of ACC and its derivatives via engineered microbial systems, thereby facilitating cost-effective development of ACC-based pharmaceuticals and agrochemicals.
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APPLIED BIOCHEMISTRY AND BIOTECHNOLOGY
·
2024
·
IF 3.1
Yuan Dongxu,Liu Bingbing,Jiang Lin,Chen Yuhuan,Xu Gang,Lin Jianping,Yang Lirong,Lian Jiazhang,Jiang Yiqi,Ye Lidan,Wu Mianbin
Xylitol is a polyol widely used in food, pharmaceuticals, and light industries. It is currently produced through the chemical catalytic hydrogenation of xylose and generates xylose mother liquor as a substantial byproduct in the procedure of xylose extraction. If xylose mother liquor could also be efficiently bioconverted to xylitol, the greenness and atom economy of xylitol production would be largely improved. However, xylose mother liquor contains a mixture of glucose, xylose, and arabinose, raising the issue of carbon catabolic repression in its utilization by microbial conversion. Targeting this challenge, the transcriptional activator XylR was overexpressed in a previously constructed xylitol-producing E. coli strain CPH. The resulting strain CPHR produced 16.61 g/L of xylitol in shake-flask cultures from the mixture of corn cob hydrolysate and xylose mother liquor (1:1, v/v) with a xylose conversion rate of 90.1%, which were 2.23 and 2.15 times higher than the starting strain, respectively. Furthermore, XylR overexpression upregulated the expression levels of xylE , xylF , xylG , and xylH genes by 2.08–2.72 times in arabinose-containing medium, suggesting the alleviation of transcriptional repression of xylose transport genes by arabinose. This work lays the foundation for xylitol bioproduction from xylose mother liquor.
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ADVANCED ENGINEERING MATERIALS
·
2024
·
IF 3.4
Zihao Song,Longjin Huang,Sheng Cui
In this work, a novel strategy for preparing Ga-doped TiO 2 composite aerogel antibacterial agent through the sol–gel method and supercritical technology is demonstrated. The specific surface area of the samples is as high as 175 m 2 g −1 and exhibits a 3D network structure with a multistage pore distribution. The obtained antibacterial agent combines the phototoxicity of TiO 2 , photo-independent toxicity of Ga 3+ , and structural properties of aerogels, showing a synergistic antibacterial effect. On the one hand, TiO 2 produces reactive oxygen species in the presence of light, thus destroying the structure of bacteria. On the other hand, Ga 3+ competitively replaces Fe 3+ and hinders the metabolic process of bacteria. Moreover, the structural properties of aerogels promote superior antimicrobial performance of antibacterial agents. Their antibacterial properties against Escherichia coli and Staphylococcus aureus bacteria are mainly investigated through the inhibition zone and natural colony counting method. In the results, it is shown that the structure and bactericidal activity of TiO 2 aerogel is effectively optimized with Ga 3+ doping. The maximum zone of inhibition of the composite aerogel against E. coli and S. aureus is 17.5 and 16 mm, and the bactericidal rate is as high as 98.57% and 96.60%, which exhibit outstanding bactericidal performance.
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Antibiotics-Basel
·
2025
·Vol. 14
·期 9
·915
·
IF 4.6
Ru Wang,Kun Mi,Aihua Lu,Chengyang Zhang,Lei Sun,Yuxiang Chen,Yuanhu Pan,Yanfei Tao,Lingli Huang
Background/Objectives: Extended-spectrum β-lactamase (ESBL)-producingEscherichia coliposes a significant global health challenge, as it leads to antimicrobial treatment failure and is associated with elevated mortality rates. The use of β-lactam/β-lactamase inhibitor combinations offers an alternative approach for combating ESBL-producing bacteria. Ceftriaxone (CRO) and sulbactam have been coadministered in the clinical settings; however, discrepancies in their pharmacokinetics raise concerns regarding the rationality of this combination.Methods: This study was designed to investigate the postβ-lactamase inhibitor effect (PLIE) under both static and dynamic conditions, with the aim of supporting the clinical application of this combination.Results: The minimum inhibitory concentration (MIC) of CRO/SBT (2:1 ratio) againstE. coliNCTC 13353 was determined to be 32/16 μg/mL. The PLIEs were determined to be −1.26, −0.57, and 0.37 h at CRO/SBT concentrations ranging from 1/2 MIC to 2 MIC, respectively. The results of CRO concentration, β-lactamase activity,blaCTX-M-15expression, and cell morphology collectively support that SBT exerts PLIEs and protects against the antibacterial activity of CRO. In the dynamic hollow-fiber infection model, CRO monotherapy showed no inhibitory effect onE. coli, whereas CRO/SBT combination therapy rapidly eliminated SBT, achieved comparable bactericidal effects, prolonged CRO exposure, and maintained low β-lactamase activity levels.Conclusions: In conclusion, CRO/SBT exerts an inhibitory effect on enzyme-producing strains by being able to produce PLIE to maintain the inhibition of β-lactamase.
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BIORESOURCE TECHNOLOGY
·
2026
·
IF 8.2
Xin Li,Haoyu Chai,Zhifan Yang,Cuiluan Ma,Yu-Cai He
Syringyl alcohol is an important lignin-derived aromatic alcohol with potential utility in the fragrance and pharmaceutical research. In this study, it was biologically synthesized from syringaldehyde. The recombinant Escherichia coli C165F-V231D expressing alcohol dehydrogenase K p ADH from Kluyveromyces polyspora enabled the effective biotransformation of syringaldehyde. Based on structural analysis and previous work for mutation of 165C to 165F and 231 V to 231D in K p ADH. This variant was created, demonstrating a markedly increased catalytic activity under low dimethyl sulfoxide (DMSO) concentrations. C165F-V231D activity could be improvemed significantly in DMSO-H 2 O (5:95, V/V). Eventually, C165F-V231D cells transformed syringaldehyde (25 mM) to syringyl alcohol in 99.7% analytical yield. This work provides a sustainable biocatalytic strategy for the synthesis of syringyl alcohol from lignin-derived aldehydes, deserves the further development lignin valorization and the green synthesis of bio-derived aromatic alcohols.
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Biotechnology Journal
·
2026
·Vol. 21
·期 2
·e70186
·
IF 3.7
Chengbin Wang,Jingxin Yin,Yali Ban,Jing Qiu,Xiaobing Yang,Chenxu Wang,Hancheng Yao,Yongjun Feng,Haijun Gao
Cis , cis -muconic acid (ccMA) is a promising platform chemical for the production of polymers and fine chemicals. However, conventional synthesis suffers from poor selectivity and environmental concerns, while plasmid-dependent biosynthesis requires expensive antibiotics and inducers for large-scale fermentation. Here, we constructed a plasmid-free and inducer-free Escherichia coli strain for the efficient production of from glucose. The biosynthetic pathway was established via the endogenous shikimate pathway, through 3-dehydroshikimate, which is sequentially converted to protocatechuate by a 3-dehydroshikimate dehydratase ( Ap AroZ R363A ), to catechol by a protocatechuate decarboxylase ( Kp AroY), and finally to ccMA by a catechol 1,2-dioxygenase ( Pp CatA). To increase the supply of ccMA, the competitive pathway genes ( aroE , pykF , and ptsG ) were disrupted, and the essential pathway genes ( galP , glK , tktA , talB , aroG , aroB , and aroD ) were overexpressed. To address the carbon flux imbalance caused by aroE and pykF knockouts, their expression was dynamically regulated using P fliA -ATG- aroE and P flgB -TTG- pykF designs. To eliminate the plasmid burden and the need for antibiotics or inducers, the ccMA pathway genes were chromosomally integrated, and the final engineered strain WMA101 produced 9.54 g/L of ccMA with a yield of 40.3% (mol/mol) from glucose in shake flasks. These studies demonstrate a sustainable, scalable platform for ccMA biomanufacturing.
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Foods
·
2025
·Vol. 14
·期 12
·2119
·
IF 5.1
Xinlei Yuan,Zihan Wang,Fang Wu,Le Cheng,Yutong Jin,Jianguo Dong,Chenyan Zheng,Yumeng Ma,Yan Jin,Bing Fang
Pesticide residues in foods can disturb the intestinal barrier and microbiota, even at a very low dose; however, studies on direct consequences on intestinal health are still lacking. Here, we evaluated the damage of imidacloprid (IMI) to the intestine and the resulting defense against enterotoxigenicEscherichia coli(ETEC) in C57BL/6J mice. After 8-week exposure to 0.06 mg /kg bodyweight/day, IMI significantly damaged intestinal structure and intestinal integrity, characterized by an increased permeability to FITC-dextran and decreased mRNA expression of tight junction proteins, as well as more broken villi and lower proportions of goblet cells and paneth cells. These were related to the suppression of the self-renewal of intestinal stem cells (ISCs), as evidenced by significantly decreased Sox9+ ISCs and increased apoptosis. Furthermore, the impaired intestinal integrity in mice exposed to low doses of IMI directly increased the susceptibility to ETEC infection and even caused death. On the other hand, exposure to 0.6 mg IMI/kg bodyweight/day lead to significantly increased contents of IL-1β and TNFα both in the intestine and serum, and significantly decreased Th1 cell and IFN-γ contents in the lamina propria during the ETEC infection. Our study suggested that the intestinal damage induced by pesticide residues would significantly decrease the defense ability of the intestine, which suggests a novel perspective when evaluating the long-term effects of food contaminates on intestinal health at low doses without significant toxicological injuries.
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