Expression of Novel L-Leucine Dehydrogenase and High-Level Production of L-Tert-Leucine Catalyzed by Engineered Escherichia coli

Jia, Yuanyuan; Xie, Yu-Li; Yang, Lulu; Shi, Hongbo; Lu, Yao; Zhang, Si-Pu; Tang, Cun-Duo; Yao, Lunguang; Kan, Yunchao

doi:10.3389/fbioe.2021.655522

Cited by 8 publications

(4 citation statements)

References 36 publications

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“…Under abiotic stress, BCAT maintains low levels of free branched-chain amino acids, which are nontoxic, to resist stress injury. 28 In the current study, BCAT activity was lower in the model group, indicating a decrease in L-leucine degradation and an increase in its levels. Therefore, L-leucine levels were higher in the model group than in the control group.…”

Section: Discussionsupporting

confidence: 41%

Modulatory Effect of Dihydromyricetin on Chronic Alcoholic Liver Injury Based on the FXR-NLRP3 Signaling Pathway and Serum and Liver Metabolomics

Wang,

Zhou,

Shi

et al. 2024

Natural Product Communications

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Background In modern diets, alcohol consumption has led to an increase in the number of cases of alcohol-related liver disease (ALD). Dihydromyricetin (DMY) is commonly used as a hepatoprotective agent owing to its remarkable efficacy in treating chronic alcoholic liver injury; however, its mechanism of action is unclear. The object of the study is to investigate the effect and mechanism of DMY in alleviating chronic ALD. Methods A mouse model of chronic ALD was established. Mice were treated with DMY for 56 days, and their biochemical parameters including liver function, blood lipids, and oxidative stress-related indices were measured. Farnesoid X receptor (FXR) expression, NOD-like receptor protein 3 (NLRP3) pathway-related protein expression, and inflammation-related gene expression were determined to elucidate the mechanism of DMY in mice with ALD. Lastly, serum and liver metabolomics-based UHPLC-Orbitrap Exploris MS analyses were used to determine the influence of the metabolism of DMY on mice with ALD. Results Pharmacodynamic studies showed that DMY could decrease aspartate transaminase, alanine transaminase, triglyceride, and low-density lipoprotein cholesterol levels, improve superoxide dismutase activity, and reduce inflammation in mice with ALD. DMY treatment protects the liver by increasing FXR protein expression and by decreasing NLRP3 pathway-related protein expression and inflammatory gene expression. Metabolomics analysis indicated that ethanol treatment mainly altered metabolism in mice. DMY could regulate 10 metabolites in serum, namely, N-α-acetyllysine, 1-pyrrolinecytosine, glutamyllysine, 5-methylcytosine, N-methylvaline, pyridoxamine, demethoxycurcumin, L-arginine, triacetin, and 15-methylpalmitate. It could also regulate 31 metabolites in the liver, including L-methionine and L-leucine. DMY treatment altered the following important pathways: valine, leucine, and isoleucine biosynthesis; cysteine and methionine metabolism; and valine, leucine, and isoleucine degradation. Correlation analyses using heatmaps revealed that the metabolic parameters are closely related to the pharmacodynamic index. Conclusion These findings indicated that DMY alleviated ALD by regulating the FXR-NLRP3 signaling pathway and could treat serum and liver metabolic disorders.

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Section: Discussionsupporting

confidence: 41%

Modulatory Effect of Dihydromyricetin on Chronic Alcoholic Liver Injury Based on the FXR-NLRP3 Signaling Pathway and Serum and Liver Metabolomics

Wang,

Zhou,

Shi

et al. 2024

Natural Product Communications

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“…Mass spectrometry conditions are as follows [ 23 ]: source: electrospray ion (ESI); source temperature: 120 ℃; mass spectrometry scan mode: positive ion; scan range: 50–1200 m/z; capillary voltage: 2.73–2.75 kV; cone voltage: 40 V; cone gas flow: 50 L h −1 ; desolvation gas flow: 600 L h −1 ; and desolvation temperature: 450 ℃. The 1 H NMR spectra were recorded on BrukerAvance DPX 250 or BrukerAvance 400 Spec-trometers.…”

Section: Methodsmentioning

confidence: 99%

Engineering Escherichia coli for high-yielding 2,5-Dimethylpyrazine synthesis from L-Threonine by reconstructing metabolic pathways and enhancing cofactors regeneration

Liu,

Wang,

Zhang

et al. 2024

Biotechnol Biofuels

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Abstract2,5-Dimethylpyrazine (2,5-DMP) is important pharmaceutical raw material and food flavoring agent. Recently, engineering microbes to produce 2,5-DMP has become an attractive alternative to chemical synthesis approach. In this study, metabolic engineering strategies were used to optimize the modified Escherichia coli BL21 (DE3) strain for efficient synthesis of 2,5-DMP using L-threonine dehydrogenase (EcTDH) from Escherichia coli BL21, NADH oxidase (EhNOX) from Enterococcus hirae, aminoacetone oxidase (ScAAO) from Streptococcus cristatus and L-threonine transporter protein (EcSstT) from Escherichia coli BL21, respectively. We further optimized the reaction conditions for synthesizing 2,5-DMP. In optimized conditions, the modified strain can convert L-threonine to obtain 2,5-DMP with a yield of 2897.30 mg/L. Therefore, the strategies used in this study contribute to the development of high-level cell factories for 2,5-DMP. Graphical Abstract

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“…Finally, the eluate containing the target protein is collected and concentrated by ultrafiltration, and the imidazole is removed, preparing the protein for further analysis or storage. 23 Enzyme Characterization. During the reaction of reductive amination, the NADH absorbance is monitored to assess enzyme activity.…”

Section: ■ Materials and Methodsmentioning

confidence: 99%

“…The target protein is then eluted using an appropriate volume of fresh elution buffer. Finally, the eluate containing the target protein is collected and concentrated by ultrafiltration, and the imidazole is removed, preparing the protein for further analysis or storage …”

Section: Methodsmentioning

confidence: 99%

Combining Flexible Region Design and Automatic Design to Enhance the Thermal Stability and Catalytic Efficiency of Leucine Dehydrogenase

Zhang,

Shi

et al. 2024

J. Agric. Food Chem.

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Leucine dehydrogenase (LeuDH, EC 1.4.1.9) can reversibly catalyze the oxidative deamination of L-leucine and some other specific α-amino acids to form the corresponding α-ketoacids. This reaction has great significance in the field of food additives and the pharmaceutical industry. The LeuDH from Exiguobacterium sibiricum (EsLeuDH) has high catalytic efficiency but limited thermal stability, hindering its widespread industrial application. In this study, a mutant N5F/I12L/A352Y of EsLeuDH (referred to as M2) was developed with enhanced thermal stability and catalytic activity through rational modification. The M2 mutant exhibits a half-life at 60 °C (t 1/2 (60 °C)) of 975.7 min and a specific activity of 69.6 U mg −1 , which is 5.4 and 2.1 times higher than those of EsLeuDH, respectively. This research may facilitate the utilization of EsLeuDH at elevated temperatures, enhancing its potential for industrial applications. The findings offer a practical and efficient approach for optimizing LeuDH and other industrial enzymes.

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Expression of Novel L-Leucine Dehydrogenase and High-Level Production of L-Tert-Leucine Catalyzed by Engineered Escherichia coli

Abstract: Graphical AbstractGenome mining of Leucine dehydrogenase and one-pot synthesis of L-tert-leucine from trimethylpyruvic acid.

Cited by 8 publications

References 36 publications

Modulatory Effect of Dihydromyricetin on Chronic Alcoholic Liver Injury Based on the FXR-NLRP3 Signaling Pathway and Serum and Liver Metabolomics

Modulatory Effect of Dihydromyricetin on Chronic Alcoholic Liver Injury Based on the FXR-NLRP3 Signaling Pathway and Serum and Liver Metabolomics

Engineering Escherichia coli for high-yielding 2,5-Dimethylpyrazine synthesis from L-Threonine by reconstructing metabolic pathways and enhancing cofactors regeneration

Combining Flexible Region Design and Automatic Design to Enhance the Thermal Stability and Catalytic Efficiency of Leucine Dehydrogenase

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