CATALYSIS LETTERS
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2025
·Vol. 155
·期 4
·1-10
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IF 2.4
Cao Songshuang,Wu Xiaomei,Ma Baodi,Xu Yi
Aliphatic chiral secondary alcohols are important chiral intermediates widely used in the synthesis of drugs and fine chemicals. The whole cells of recombinant E. coli co-expressing alcohol dehydrogenase and glucose dehydrogenase were used to catalyze the asymmetric reduction of three fatty ketones without the addition of exogenous cofactors. The reaction medium and reaction conditions were systematically studied. As expected, the kinds of co-solvents, the molar ratio of glucose/substrate, temperature, pH and cell amount have important influence on the reaction. Under the optimal condition (glucose at 1.5 times substrate concentration, isopropanol at 2%, PBS (0.1 M, pH 8.0), and 45℃), the cells co-expressing both enzymes efficiently catalyzed the asymmetric reduction of three fatty ketones to the corresponding ( S )-fatty alcohols. The conversion and ee value of the products were both greater than 99% after 8 h reaction at 1 M of substrate concentration, and the space-time yield was between 346 ~ 470 g/(L·d). All the products can be obtained on gram scale. The results indicated that this method has potential application value in the efficient and green synthesis of aliphatic chiral secondary alcohols. Graphical
研究领域:
生物化学
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合成生物学
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高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
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2018
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IF 5.667
Shuyun Zhu,Cuihua Lei,Jing Sun,Xian-En Zhao,Xiao Wang,Xiaolu Yan,Wei Liu,Hua Wang
In the present study, a novel colorimetric sensing platform was constructed for probing the NAD + /NADH-dependent biocatalytic transformations. Manganese dioxide (MnO 2 ) nanosheets as an oxidase-mimicking nanomaterial could directly oxidize 3,3′,5,5′-tetramethylbenzidine (TMB) into oxTMB without the need of H 2 O 2 . Importantly, it was found that NADH could easily trigger the decomposition of MnO 2 nanosheets, causing the decrease of solution absorbance. This is demonstrated by the enzyme catalysis reactions of two dehydrogenase models, ethanol dehydrogenase (ADH) and lactate dehydrogenase (LDH), where NADH is formed and consumed, respectively, in their physiological enzymatic reactions. A MnO 2 -TMB sensing platform was thereby proposed for the analysis of their catalytic reaction substrates including ethanol and pyruvic acid. The detection limits of ethanol and pyruvic acid can reach 5.0 μM and 100 nM, respectively. The developed optical biosensing platform can be used to develop other dehydrogenase-based biosensors followed by monitoring their substrates.
研究领域:
生物化学
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纳米生物学
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分析化学
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高影响因子
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
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2020
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IF 4.192
Tian Li,Guo-Song Liu,Wei Zhou,Min Jiang,Yu-Hong Ren,Xin-Yi Tao,Min Liu,Ming Zhao,Feng-Qing Wang,Bei Gao,Dong-Zhi Wei
Squalene has wide applications in the food and pharmaceutical industries. Engineering microbes to produce squalene is a promising alternative for traditional production approaches. In this study, squalene production was enhanced to 978.24 mg/L through stepwise overexpression of the enzymes that catalyze acetyl-CoA to squalene. Subsequently, to increase the activity of HMG-CoA reductase and alleviate the high dependence on NADPH, the HMG-CoA reductase (NADH-HMGR) from Silicibacter pomeroyi, highly specific for NADH, was introduced, which increased squalene production to 1086.31 mg/L. Native ethanol dehydrogenase ADH2 and acetaldehyde dehydrogenase ADA from Dickeya zeae were further overexpressed, which enhanced the capability to utilize ethanol for squalene synthesis and endowed the engineered strain with greater adaptability to high ethanol concentrations. Finally, a remarkable squalene production of 9472 mg/L was obtained from ethanol via carbon source-controlled fed-batch fermentation. This study will greatly accelerate the process of developing microbial cell factories for squalene production.
研究领域:
生物技术
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代谢工程
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微生物生物技术
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DOI
PubMed
高影响因子
ORGANIC PROCESS RESEARCH & DEVELOPMENT
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2022
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IF 3.858
Ling He,Wenjie Ye,Youyu Xie,Qinghai Liu,Hualei Wang,Dongzhi Wei
Bioreductions catalyzed by ketoreductases play an important role in the synthesis of chiral alcohols. However, the synthesis of (R)-2-chloro-1-(3,4-difluorophenyl)ethanol (24b), an important chiral intermediate for the synthesis of the anticoagulant ticagrelor, poses significant challenges in terms of high substrate concentration requirements that limit its production. In this study, a novel NADH-dependent, short-chain dehydrogenase, PpKR8, from Paraburkholderia phymatum STM815, exhibited excellent enantioselectivity and high activity for the production of (R)-CFPL (24b) through the reduction of 2-chloro-1-(3,4-difluorophenyl)ethanone (24a). The coexpression of PpKR8 and glucose dehydrogenase from Bacillus subtilis in Escherichia coli allowed up to 300 g/L (1.57 M) CFPO (24a) to be completely converted into 24b with 99.9% enantiomeric excess and a high space–time yield (728 g/(L day)). Substrate specificity assays demonstrated a broad substrate spectrum for PpKR8, which included 35 α/β-ketoesters, aromatic ketones, and heterocyclic ketones. Moreover, three additional optically pure chiral alcohols that are used as important drug intermediates were synthesized at high substrate concentrations (150–330 g/L), demonstrating the excellent industrial potential of PpKR8-mediated bioreductions.
研究领域:
生物化学
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酶工程
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制药工业
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DOI
高影响因子
Journal of Biological Research-Thessaloniki
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2020
·Vol. 27
·期 1
·1-10
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IF 2.121
Cao Yujin,Mu Hui,Guo Jing,Liu Hui,Zhang Rubing,Liu Wei,Xian Mo,Liu Huizhou
Background The fuel ethanol industry has made tremendous progress in the last decades. Ethanol can be obtained by fermentation using a variety of biomass materials as the feedstocks. However, few studies have been conducted on ethanol utilization by microorganisms. The price of petroleum-derived ethanol, easily made by the hydrolysis of ethylene, is even lower than that of bioethanol. If ethanol can be metabolized by microorganisms to produce value-added chemicals, it will open a new door for the utilization of inexpensive ethanol resources.Results We constructed an engineered Escherichia coli strain which could utilize ethanol as the sole carbon source. The alcohol dehydrogenase and aldehyde dehydrogenase from Aspergillus nidulans was introduced into E. coli and the recombinant strain acquired the ability to grow on ethanol. Cell growth continued when ethanol was supplied after glucose starvation and 2.24 g L−1 of ethanol was further consumed during the shake-flasks fermentation process. Then ethanol was further used for the production of mevalonic acid by heterologously expressing its biosynthetic pathway. Deuterium-labeled ethanol-D6 as the feedstock confirmed that mevalonic acid was synthesized from ethanol.Conclusion sThis study demonstrated the possibility of using ethanol as the carbon source by engineered E. coli strains. It can serve as the basis for the construction of more robust strains in the future though the catabolic capacity of ethanol should be further improved.
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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PubMed
高影响因子
Microbial Biotechnology
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2020
·Vol. 14
·期 2
·444-452
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IF 5.328
Meng-Nan Han,Xu-Ming Wang,Chao-Hong Pei,Chao Zhang,Zhidong Xu,Hong-Lei Zhang,Wei Li
Summary Chiral aromatic alcohols have received much attention due to their widespread use in pharmaceutical industries. In the asymmetric synthesis processes, the excellent performance of alcohol dehydrogenase makes it a good choice for biocatalysts. In this study, a novel and robust medium-chain alcohol dehydrogenase RhADH from Rhodococcus R6 was discovered and used to catalyse the asymmetric reduction of aromatic ketones to chiral aromatic alcohols. The reduction of 2-hydroxyacetophenone (2-HAP) to ( R )-(-)-1-phenyl-1,2-ethanediol (( R )-PED) was chosen as a template to evaluate its catalytic activity. A specific activity of 110 U mg −1 and a 99% purity of e.e . was achieved in the presence of NADH. An efficient bienzyme-coupled catalytic system (RhADH and formate dehydrogenase, CpFDH) was established using a two-phase strategy (dibutyl phthalate and buffer), which highly raised the tolerated substrate concentration (60 g l −1 ). Besides, a broad range of aromatic ketones were enantioselectively reduced to the corresponding chiral alcohols by this enzyme system with highly enantioselectivity. This system is of the potential to be applied at a commercial scale.
研究领域:
生物化学
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生物技术
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药物化学
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PubMed
高影响因子
PREPARATIVE BIOCHEMISTRY & BIOTECHNOLOGY
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2019
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IF 1.117
Chuhong Zong,Xu Zhang,Fei Yang,Yafeng Zhou,Nan Chen,Zuisu Yang,Guofang Ding,Fangmiao Yu,Yunping Tang
(S)-1-(2, 6-dichloro-3-fluorophenyl) ethanol, the key chiral intermediate of crizotinib, was prepared from 1-(2, 6-dichloro-3-fluorophenyl) ethanone using the alcohol dehydrogenases from Lactobacillus kefir (ADH-LK) with a tetrad mutant (ADH-LKM, F147L/Y190P/V196L/A202W), coupled with glucose dehydrogenase (GDH). In the present study, ADH-LKM and GDH were successfully heterologous expressed in recombinant Escherichia coli. During the regeneration of NADPH with GDH, 150 g/L substrate was totally transformed into target chiral alcohol with an enantiomeric excess value of 99.9% after 12 h at 30 °C (pH 7.0). Our study demonstrates the potential for industrial green production of the key chiral intermediate of crizotinib.
研究领域:
生物催化
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药物化学
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酶工程
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DOI
PubMed
高影响因子
Advanced Science
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2024
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IF 14.3
Likun Luan,Yingfang Zhang,Xiuling Ji,Boxia Guo,Shaoyu Song,Yuhong Huang,Suojiang Zhang
Multi-enzymatic cascade reaction provides a new avenue for C─C coupling directly from CO 2 under mild conditions. In this study, a new pathway with four enzymes including formate dehydrogenase (PaFDH), formaldehyde dehydrogenase (BmFADH), glycolaldehyde synthase (PpGALS), and alcohol dehydrogenase (GoADH) is developed for directly converting CO 2 gas molecules to ethylene glycol (EG) in vitro. A rhodium-based NADH regeneration electrode is constructed to continuously provide the proton and electron of this multi-enzymatic cascade reaction. The prepared electrode can reach the Faradaic Efficiency (FE) of 82.9% at −0.6 V (vs. Ag/AgCl) and the NADH productivity of 0.737 mM h −1 . Shortening the reaction path is crucial for multi-enzymatic cascade reactions. Here, a hydrogen-bonded organic framework (HOF) nano-reactor is successfully developed to immobilize four enzymes in one pot with a striking enzyme loading capacity (990 mg enzyme g −1 material). Through integrating and optimization of NADH electro-regeneration and enzymatic catalysis in one pot, 0.15 mM EG is achieved with an average conversion rate of 7.15 × 10 −7 mmol CO 2 min −1 mg −1 enzymes in 6 h. These results shed light on electro-driven multi-enzymatic cascade conversion of C─C coupling from CO 2 in the nano-reactor.
研究领域:
生物化学
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生物技术
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电化学
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酶工程
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纳米技术
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DOI
PubMed
高影响因子
BIOCHEMICAL ENGINEERING JOURNAL
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2024
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IF 3.7
Shu-Ping Zou,Wei Ding,Yu-Yue Han,Kun Niu,Ya-Ping Xue,Yu-Guo Zheng
L-Phosphinothricin (L-PPT) is the active compound of a broad-spectrum herbicide. In this study, we biosynthesized L-PPT via the asymmetric reduction of 2-carboxy-4-(hydroxymethylphosphoryl) butyric acid (PPO) using a whole-cell biocatalyst. Glutamate dehydrogenase from Pseudomonas moorei ( Pm GluDH), Glucose dehydrogenase from Exiguobacterium sibiricum ( Es GDH) and NAD kinase from Corynebacterium glutamicum ( Cg NADK) were coexpressed in E. coli , significantly increasing the catalytic efficiency by regenerating more NADPH to promote PPO reduction. Following the optimization of balancing the co-expression of the three enzymes in pETDuet-1 and pCDFDuet-1, the coexpressing strain increases in the intracellular NADP(H) concentration to 54.9 μmol/g DCW, which was 3.8-fold higher than the control. The co-expression system exhibited optimal activity at a temperature of 35°C, pH 7.5, and the glucose to PPO molar ratio of 1.4:1. In a 30 L bioconversion system the final concentration and productivity of L-PPT reached 79.8 g/L (>99% ee ) and 1440 g/(L·d), respectively. These results will contribute to the industrial biological production of L-PPT.
研究领域:
生物技术
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生化工程
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代谢工程
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高影响因子
ACS Applied Nano Materials
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2025
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IF 5.5
Xintong Dong,Mengyuan Li,Xueyan Zhong,Mingqi Sun,Xiaoying Zhang,Ying Wang,Yao Wang,Ying Guo
Electrochemical biosensors based on nicotinamide adenine dinucleotide (NADH)-dependent dehydrogenases hold great potential in analytical applications. However, previous biosensors can lead to enzyme detachment and inactivation, thus limiting their applications. This study presents a novel sensing platform based on a three-dimensional MXene-graphene oxide sponge (3D MG). The nanoporous sponge was synthesized by polyethylene glycol (PEG) assisted cross-linking of MXene and small-sized graphene oxide (GO) nanosheets, thereby forming a high-strength porous conductive network and addressing the instability of enzyme-based biosensors. This sponge significantly enhances enzyme immobilization (reducing leaching by 62.27% compared to MXene films) and maintains enzymatic activity under extreme conditions. The 3D MG sensing platform was further fabricated into two biosensors by immobilizing NADH-dependent dehydrogenases (with glutamate dehydrogenase and formate dehydrogenase employed as model enzymes, respectively), using the (2,2′-bipyridyl) (pentamethylcyclopentadienyl)-rhodium complex ([Cp*Rh(bpy)Cl]+) as a mediator that enables the regeneration of bioactive NADH. The two biosensors exhibited low detection limits (0.332 μM for NH4+ and 0.188 mM for HCO3–), excellent reproducibility (relative standard deviation < 4%), and long-term stability (94.82% and 92.63% signal retention after 60 days). The fabrication of 3D MG with a robust nanoporous structure and an enzyme-friendly environment provides a strategy for designing long-life multifunctional enzyme-based biosensors.
研究领域:
生物传感器
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纳米材料
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电化学
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酶工程
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分析化学
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材料科学
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高影响因子
BIOCHEMICAL ENGINEERING JOURNAL
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2022
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IF 4.446
Shuping Zou,Zhentao Jiang,Heng Tang,Feng Cheng,Yaping Xue,Yuguo Zheng
The enzymatic asymmetric synthesis of L -phosphinothricin ( L -PPT) from 2-carboxyl-4-(hydroxymethylphosporyl) butyric acid (PPO) is a green and efficient method. In our previous works, we constructed a glutamate dehydrogenase (GluDH) and glucose dehydrogenase (GDH) co-expression strain, Escherichia coli BL21 (DE3)/pETDuet- gludh-gdh , which can asymmetrically reduce PPO to L -PPT. To improve the stability and recyclability of the biocatalyst , we developed a novel method for whole-cell immobilization with cross-linked cell aggregates (CLCAs) and employed immobilized cells in L -PPT production. Montmorillonite was used as a carrier, and dextran polyaldehyde (DPA) was developed as a new cross-linker for cell immobilization . The highest activity recovery (79.2%) was obtained under the following conditions: 20 g/L cell concentration, 0.9% montmorillonite concentration, 1.2% polyethyleneimine (PEI) concentration, and 1.2% DPA concentration. The half-lives of CLCAs were 169.10 h (35 °C), 17.39 h (45 °C), and 10.65 h (55 °C), which were 1.87-, 3.70-, and 10.78-fold than that of free cells, respectively. The CLCAs showed good reusability, retaining over 85% of their initial activities after 10 cycles of utilization. The total yield of L -PPT reached 667 mmol/g cells, and the enantiomeric excess value was more than 99%. The immobilized whole-cell in the form of CLCAs is a valuable biocatalyst for industrial applications.
研究领域:
生物化学
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工业微生物学
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高影响因子
ENZYME AND MICROBIAL TECHNOLOGY
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2022
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IF 3.705
Jinbin Liu,Kaipeng Wang,Mian Wang,Huaxiang Deng,Xiaodong Chen,Yueling Shang,Xiaochen Liu,Xiaohong Yu
An engineered Escherichia coli was constructed by co-expressing L -amino acid deaminase, α-keto acid decarboxylase, alcohol dehydrogenase , and glucose dehydrogenase through two plasmids for tyrosol production. The activity of the rate-limiting enzyme L -amino acid deaminase from Cosenzaea myxofaciens (CmAAD) toward tyrosine was improved by structure-guided modification. The enzyme activity of triple mutant CmAAD V438G/K147V/R151E toward tyrosine was ~5.12-fold higher than that of the wild-type CmAAD. Secondly, the plasmid copy numbers and the gene orders were optimized to improve the titer of tyrosol. Finally, the recombinant strain CS-6 transformed 10 mM tyrosine into 9.56 ± 0.64 mM tyrosol at 45 ℃, and the space-time yield reached 0.478 mM·L −1 ·h −1 . This study proposes a novel idea for the efficient and natural production of tyrosol, which has great potential for industrial application.
研究领域:
生物技术
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酶学
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微生物学
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DOI
PubMed
高影响因子
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