摘要：
Orcinol glucoside (OG), mainly found in the rhizome of the traditional Chinese herb Curculigo orchioides Gaertn, is noted for its antidepressant effects. In this study, an efficient screening pipeline was established for identifying the highly active orcinol synthase (ORS) and UDP-dependent glycosyltransferase (UGT) involved in the biosynthesis of OG by combining transcriptome analysis, structure-based virtual screening, and in vitro enzyme activity assays. By enhancing the downstream pathway, metabolic engineering and fermentation optimization, the OG production in Yarrowia lipolytica was improved 100-fold, resulting in a final yield of 43.46 g/L (0.84 g/g DCW), which is almost 6,400-fold higher than the extraction yield from C. orchioides roots. This study provides a reference for rapid identification of functional genes and high-yield production of natural products.
摘要：
Recently, nanomaterial lubricant additives have attracted the attention of tribologists. Herein, a novel cobalt phosphate octahydrates (CoP) nanoflower was synthesized, and as an oil-base additive, CoP exhibited extraordinary tribological performance for titanium alloy. Compared with the pure PAO8, the friction coefficient (COF) and wear rate (WR) has been reduced by 69.77% and 99.27% at a concentration of 10 wt% CoP. Besides, SEM, EDS, and XPS characterization confirmed that CoP could fill and repair the surface of titanium alloy to make the worn surface smoother, and be deposited on the surface to form a deposition layer to protect the titanium alloy from severe wear. The low shear force between CoP nanosheets in the deposition layer promotes lubrication, achieving low COF.
期刊：
World Journal of Microbiology and Biotechnology,2023年39(3):1-12 ISSN：0959-3993
通讯作者：
Xiaomin Hu<&wdkj&>Hairong Xiong
作者机构：
[Wang, Yawei] Wuhan Polytech Univ, Coll Life Sci & Technol, Wuhan 430048, Peoples R China.;[Xiong, Hairong; Hu, Xiaomin; Zhao, Puying] South Cent Univ Nationalities, Coll Life Sci, Wuhan 430074, Peoples R China.;[Zhou, Ying] Wuhan Sunhy Biol Co Ltd, Wuhan 430205, Peoples R China.
通讯机构：
[Xiaomin Hu; Hairong Xiong] C;College of Life Science, South-Central University for Nationalities, Wuhan, China<&wdkj&>College of Life Science, South-Central University for Nationalities, Wuhan, China
摘要：
Protein hydrolysates are easily digested and utilized by humans and animals, and are less likely to cause allergies. Protein hydrolysis caused by endopeptidases often leads to the exposure of hydrophobic amino acids at the ends of peptides, which consequently causes bitter taste. Microbial aminopeptidases remove the exposed hydrophobic amino acids at the ends of aminopeptides, which improves taste, allowing for easier production. This processe is attacking significant attention from industry and laboratories. Aminopeptidases selectively hydrolyze peptide bonds from the N-terminal of proteins or peptides to produce free amino acids. Aminopeptidases can be classified into leucine, lysine, methionine and proline aminopeptidases by hydrolyzed N-terminal residues; metallo-, serine- and cysteine- aminopeptidases by the reaction mechanisms; dipeptide and triphoptide enzymes by the released number of amino acid residues at the end of hydrolyzed peptides; or acidic, neutral and basic aminopeptidases by their optimal hydrolysis pH. Commercial aminopeptidases are generally produced by microbial fermentation, and are mainly applied in the debittering of protein hydrolysates, the deep hydrolysis of protein, and the production of condiments, cheese, and bioactive peptides, as well as for disease detection in the medical industry.
通讯机构：
[Limei Wang] C;College of Life Science and Technology, Wuhan Polytechnic University, Wuhan 430023, China<&wdkj&>Author to whom correspondence should be addressed.
关键词：
cell pathway;corilagin from Euryale ferox Salisb shell;inflammatory reaction;Raw264.7 macrophage
摘要：
(1) Background: Euryale ferox Salisb is a large aquatic plant of the water lily family and an edible economic crop with medicinal value. The annual output of Euryale ferox Salisb shell in China is higher than 1000 tons, often as waste or used as fuel, resulting in waste of resources and environmental pollution. We isolated and identified the corilagin monomer from Euryale ferox Salisb shell and discovered its potential anti-inflammatory effects. This study aimed to investigate the anti-inflammatory effect of corilagin isolated from Euryale ferox Salisb shell. (2) Methods: We predict the anti-inflammatory mechanism by pharmacology. LPS was added to 264.7 cell medium to induce an inflammatory state, and the safe action range of corilagin was screened using CCK-8. The Griess method was used to determine NO content. The presence of TNF-alpha, IL-6, IL-1 beta, and IL-10 was determined by ELISA to evaluate the effect of corilagin on the secretion of inflammatory factors, while that of reactive oxygen species was detected by flow cytometry. The gene expression levels of TNF-alpha, IL-6, COX-2, and iNOS were determined using qRT-PCR. qRT-PCR and Western blot were used to detect the mRNA and expression of target genes in the network pharmacologic prediction pathway. (3) Results: Network pharmacology analysis revealed that the anti-inflammatory effect of corilagin may be related to MAPK and TOLL-like receptor signaling pathways. The results demonstrated the presence of an anti-inflammatory effect, as indicated by the reduction in the level of NO, TNF-alpha, IL-6, IL-1 beta, IL-10, and ROS in Raw264.7 cells induced by LPS. The results suggest that corilagin reduced the expression of TNF-alpha, IL-6, COX-2, and iNOS genes in Raw264.7 cells induced by LPS. The downregulation of the phosphorylation of I kappa B-alpha protein related to the toll-like receptor signaling pathway and upregulation of the phosphorylation of key proteins in the MAPK signaling pathway, P65 and JNK, resulted in reduced tolerance toward lipopolysaccharide, allowing for the exertion of the immune response. (4) Conclusions: The results demonstrate the significant anti-inflammatory effect of corilagin from Euryale ferox Salisb shell. This compound regulates the tolerance state of macrophages toward lipopolysaccharide through the NF-kappa B signaling pathway and plays an immunoregulatory role. The compound also regulates the expression of iNOS through the MAPK signaling pathway, thereby alleviating the cell damage caused by excessive NO release.
摘要：
Zearalenone (ZEN) is one of the most prevalent estrogenic mycotoxins, is produced mainly by the Fusarium family of fungi, and poses a risk to the health of animals. Zearalenone hydrolase (ZHD) is an important enzyme capable of degrading ZEN into a non-toxic compound. Although previous research has investigated the catalytic mechanism of ZHD, information on its dynamic interaction with ZEN remains unknown. This study aimed to develop a pipeline for identifying the allosteric pathway of ZHD. Using an identity analysis, we identified hub genes whose sequences can generalize a set of sequences in a protein family. We then utilized a neural relational inference (NRI) model to identify the allosteric pathway of the protein throughout the entire molecular dynamics simulation. The production run lasted 1 microsecond, and we analyzed residues 139-222 for the allosteric pathway using the NRI model. We found that the cap domain of the protein opened up during catalysis, resembling a hemostatic tape. We used umbrella sampling to simulate the dynamic docking phase of the ligand-protein complex and found that the protein took on a square sandwich shape. Our energy analysis, using both molecular mechanics/Poisson-Boltzmann (Generalized-Born) surface area (MMPBSA) and Potential Mean Force (PMF) analysis, showed discrepancies, with scores of -8.45 kcal/mol and -1.95 kcal/mol, respectively. MMPBSA, however, obtained a similar score to that of a previous report.
摘要：
Combined with the Konjac transcriptome database of our laboratory and internal reference genes commonly used in plants, the eight candidate internal reference genes were screened and detected. They are the 25S ribosomal RNA gene (25S rRNA), 18S ribosomal RNA gene (18S rRNA), actin gene (ACT), glyceraldehyde-3-phosphate dehydrogenase gene (GAPDH), ubiquitin gene (UBQ), beta-tubulin gene (beta-TUB), eukaryotic elongation factor 1-ffgene(eEF-1 alpha), and eukaryotic translation initiation factor 4 beta-1 gene (eIF-4 alpha). The results of GeNorm, Normfinder, and BestKeeper were analyzed comprehensively. The data showed that the expression levels of 25S rRNA, 18S rRNA, and ACT at the reproductive periods, eEF-1 alpha and eIF-4 alpha at the nutritional periods, and eEF-1 alpha, UBQ, and ACT at different leaf developmental periods were stable. These identified and stable internal reference genes will provide the basis for the subsequent molecular biology-related studies of Konjac.
作者机构：
[Bao, Weiwei; Yang, Shihui; Peng, Qiqun] Hubei Univ, Sch Life Sci, State Key Lab Biocatalysis & Enzyme Engn, Wuhan 430062, Peoples R China.;[Shen, Wei] Wuhan Polytech Univ, Sch Life & Technol, Wuhan 430023, Peoples R China.;[Du, Jun] China Biotech Fermentat Ind Assoc, Beijing 100833, Peoples R China.
通讯机构：
[Wei Shen; Shihui Yang] A;Authors to whom correspondence should be addressed.<&wdkj&>School of Life and Technology, Wuhan Polytechnic University, Wuhan 430023, China<&wdkj&>Authors to whom correspondence should be addressed.<&wdkj&>State Key Laboratory of Biocatalysis and Enzyme Engineering, School of Life Sciences, Hubei University, Wuhan 430062, China
摘要：
Biorefinery to produce value-added biochemicals offers a promising alternative to meet our sustainable energy and environmental goals. Acetoin is widely used in the food and cosmetic industries as taste and fragrance enhancer. The generally regarded as safe (GRAS) bacterium Zymomonas mobilis produces acetoin as an extracellular product under aerobic conditions. In this study, metabolic engineering strategies were applied including redistributing the carbon flux to acetoin and manipulating the NADH levels. To improve the acetoin level, a heterologous acetoin pathway was first introduced into Z. mobilis, which contained genes encoding acetolactate synthase (Als) and acetolactate decarboxylase (AldC) driven by a strong native promoter Pgap. Then a gene encoding water-forming NADH oxidase (NoxE) was introduced for NADH cofactor balance. The recombinant Z. mobilis strain containing both an artificial acetoin operon and the noxE greatly enhanced acetoin production with maximum titer reaching 8.8 g/L and the productivity of 0.34 g center dot L-1 center dot h(-1). In addition, the strategies to delete ndh gene for redox balance by native I-F CRISPR-Cas system and to redirect carbon from ethanol production to acetoin biosynthesis through a dcas12a-based CRISPRi system targeting pdc gene laid a foundation to help construct an acetoin producer in the future. This study thus provides an informative strategy and method to harness the NADH levels for biorefinery and synthetic biology studies in Z. mobilis.
摘要：
Zymomonas mobilis is an important industrial bacterium ideal for biorefinery and synthetic biology studies. High-throughput CRISPR-based genome editing technologies have been developed to enable targeted engineering of genes and hence metabolic pathways in the model ZM4 strain, expediting the exploitation of this biofuel-producing strain as a cell factory for sustainable chemicals, proteins and biofuels production. As these technologies mainly take plasmid-based strategies, their applications would be impeded due to the fact that curing of the extremely stable plasmids is laborious and inefficient. Whilst counterselection markers have been proven to be efficient for plasmid curing, hitherto only very few counterselection markers have been available for Z. mobilis. We constructed a conditional lethal mutant of the pheS gene of Z. mobilis ZM4, clmPheS, containing T263A and A318G substitutions and coding for a mutated alpha-subunit of phenylalanyl-tRNA synthetase to allow for the incorporation of a toxic analog of phenylalanine, p-chloro-phenylalanine (4-CP), into proteins, and hence leading to inhibition of cell growth. We demonstrated that expression of clmPheS driven by a strong Pgap promoter from a plasmid could render the Z. mobilis ZM4 cells sufficient sensitivity to 4-CP. The clmPheS-expressing cells were assayed to be extremely sensitive to 0.2 mM 4-CP. Subsequently, the clmPheS-assisted counterselection endowed fast curing of genome engineering plasmids immediately after obtaining the desired mutants, shortening the time of every two rounds of multiplex chromosome editing by at least 9 days, and enabled the development of a strategy for scarless modification of the native Z. mobilis ZM4 plasmids. This study developed a strategy, coupling an endogenous CRISPR-based genome editing toolkit with a counterselection marker created here, for rapid and efficient multi-round multiplex editing of the chromosome, as well as scarless modification of the native plasmids, providing an improved genome engineering toolkit for Z. mobilis and an important reference to develope similar genetic manipulation systems in other non-model organisms.
通讯机构：
[Zhiguo Liu] H;Hubei Province Engineering Research Center of Healthy Food, School of Biology and Pharmaceutical Engineering, Wuhan Polytechnic University, Wuhan 430023, China
摘要：
Scope: Circadian disorder and high-fat diet (HFD) can disturb lipid metabolism homeostasis and may promote the development of various metabolic diseases. The relationship between them is of great concern. This study aimed to explore the effects of Per1/Per2 double knockout (DKO) on hepatic lipid metabolism in mice under HFD and HFD with docosahexaenoic acid (DHA) substitution. Methods and results: Both wild type (WT) and DKO male C57BL/6 mice were fed with normal chow diet (CON), HFD, or HFD with DHA substitution (AO) for 15 weeks. At the end of the experiment, mice were sacrificed at zeitgeber time (ZT) 0 (7:00 am) or ZT12 (7:00 pm). Pathological indicators were determined using histological and biochemical methods. Hepatic transcriptome sequencing analysis showed that DKO mice exhibited multiple dysfunctions in diurnal rhythm, drug metabolism, cell cycle, cancer pathways, and lipid metabolism. HFD had greater effects on fatty acid oxidation and cholesterol synthesis and metabolism in Per1-/-Per2-/- mice, which was improved by DHA substitution. Conclusions: Per1/Per2 played an important role in the circadian regulation of hepatic lipid metabolism, and DKO mice were more sensitive to HFD. DHA can improve circadian-related lipid metabolism disruption induced by HFD in mice.(c) 2023 Beijing Academy of Food Sciences. Publishing services by Elsevier B.V. on behalf of KeAi Communications Co., Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
期刊：
Ecotoxicology and Environmental Safety,2022年240:113698 ISSN：0147-6513
通讯作者：
Xiaochun Guo<&wdkj&>Dongru Qiu
作者机构：
[Liu, Ying; Wan, Zhengfen; Guo, Xiaochun; Lu, Shaoyong; Liu, Xiaohui] Chinese Res Inst Environm Sci, State Key Lab Environm Criteria & Risk Assessment, Natl Engn Lab Lake Pollut Control & Ecol Restorat, Beijing 100012, Peoples R China.;[Liu, Xiaohui] Tsinghua Univ, Sch Environm, Beijing 100084, Peoples R China.;[Chen, Jing] Wuhan Polytech Univ, Coll Life Sci & Technol, Wuhan 430070, Peoples R China.;[Chen, Jing; Qiu, Dongru] Chinese Acad Sci, Inst Hydrobiol, Wuhan 430072, Peoples R China.
通讯机构：
[Xiaochun Guo] S;[Dongru Qiu] I;Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072, China<&wdkj&>State Key Laboratory of Environmental Criteria and Risk Assessment, National Engineering Laboratory for Lake Pollution Control and Ecological Restoration, Chinese Research Academy of Environmental Sciences, Beijing 100012, China
摘要：
Feruloyl esterase is a subclass of α/β hydrolase, which could release ferulic acid from biomass residues for use as an efficient additive in food or pharmaceutical industries. In the present study, a feruloyl esterase with broad substrate specificity was characterised and secreted by Bacillus subtilis WB600. After codon usage optimisation and signal peptide library screening, the secretion amount of feruloyl esterase was enhanced by up to 10.2-fold in comparison with the base strain. The site-specific amino acid substitutions that facilitate protein folding further improved the secretion by about 1.5-fold. The purified rationally designed enzyme exhibited maximal activity against methyl ferulate at pH 6.5 and 65 °C. In the solid-state fermentation, the genetically engineered B. subtilis released about 37% of the total alkali-extractable ferulic acid in maize bran. This study provides a promising candidate for ferulic acid production and demonstrates that the secretion of a heterologous enzyme from B. subtilis can be cumulatively improved by changes in protein sequence features.
通讯机构：
[Liu, Z.] H;Hubei Province Engineering Research Center of Healthy Food, School of Biology and Pharmaceutical Engineering, Wuhan Polytechnic University, Wuhan, China
关键词：
bile acid;circadian rhythm;high-fat diet;intestinal flora
通讯机构：
[Min, T ] W;Wuhan Polytech Univ, Coll Food Sci & Engn, Wuhan 430023, Peoples R China.
关键词：
Fresh-cut Chinese water chestnut;melatonin;storage quality;phenolic metabolism;reactive oxygen species metabolism
摘要：
Fresh-cut Chinese water chestnuts (CWCs) are prone to quality deterioration during storage, which does not meet consumer demand. In this study, the effect of exogenous melatonin (5 mmol center dot L-1) on the quality and potential mechanisms in fresh-cut CWC was investigated. The results showed that melatonin treatment alleviated the cut-surface discoloration of CWCs. Not only did this treatment significantly slow down the increase in browning degree and yellowness (b*) as well as the decrease in lightness (L*), but it also significantly delayed the loss of weight and total soluble solids. Further investigations indicated that melatonin-treated fresh-cut CWCs exhibited significantly lower total phenolics and soluble quinones and suppressed the activities of phenylalanine ammonia-lyase, polyphenol oxidase, and peroxidase. Meanwhile, when fresh-cut CWCs were treated with melatonin, the total flavonoid concentration was significantly decreased compared to the control. Additionally, melatonin significantly inhibited the accumulation of H2O2 and malondialdehyde as well as enhanced the activities of superoxide dismutase and catalase by promoting the production of O-2(-center dot). In summary, melatonin treatment may delay the surface discoloration of fresh-cut CWCs by inhibiting phenolic compound metabolism and improving antioxidant capacity, thereby effectively maintaining the quality and prolonging the shelf life of fresh-cut CWCs.
作者机构：
[Hao, Yile; Zheng, Yanli] Wuhan Polytech Univ, Coll Life Sci & Technol, Wuhan 430023, Peoples R China.;[Hao, Yile; Yang, Shihui; Peng, Wenfang; Li, Jie; Wang, Qinhua] Hubei Univ, Hubei Engn Res Ctr Bioenzyme Catalysis,Sch Life S, Environm Microbial Technol Ctr Hubei Prov,State K, Hubei Collaborat Innovat Ctr Green Transformat Bi, Wuhan 430062, Peoples R China.
通讯机构：
[Zheng, Yanli] W;[Peng, Wenfang] H;Wuhan Polytech Univ, Coll Life Sci & Technol, Wuhan 430023, Peoples R China.;Hubei Univ, Hubei Engn Res Ctr Bioenzyme Catalysis,Sch Life S, Environm Microbial Technol Ctr Hubei Prov,State K, Hubei Collaborat Innovat Ctr Green Transformat Bi, Wuhan 430062, Peoples R China.
关键词：
large genomic fragments deletion;high-efficiency;genome editing;CRISPR-Cas;Cas3 nickase
摘要：
New CRISPR-based genome editing technologies are developed to continually drive advances in life sciences, which, however, are predominantly derived from systems of Type II CRISPR-Cas9 and Type V CRISPR-Cas12a for eukaryotes. Here we report a novel CRISPR-n(nickase)Cas3 genome editing tool established upon a Type I-F system. We demonstrate that nCas3 variants can be created by alanine-substituting any catalytic residue of the Cas3 helicase domain. While nCas3 overproduction via plasmid shows severe cytotoxicity, an in situ nCas3 introduces targeted double-strand breaks, facilitating genome editing without visible cell killing. By harnessing this CRISPR-nCas3 in situ gene insertion, nucleotide substitution and deletion of genes or genomic DNA stretches can be consistently accomplished with near-100% efficiencies, including simultaneous removal of two large genomic fragments. Our work describes the first establishment of a CRISPR-nCas3-based genome editing technology, thereby offering a simple, yet useful approach to convert the naturally most abundantly occurring Type I systems into advanced genome editing tools to facilitate high-throughput prokaryotic engineering.
通讯机构：
[Wenfu Hou] C;[Wen Wang] S;College of Food Science and Engineering, Wuhan Polytechnic University, Wuhan, China<&wdkj&>State Key Laboratory for Managing Biotic and Chemical Threats to the Quality and Safety of Agro-products, MOA Laboratory of Quality & Safety Risk Assessment for Agro-products (Hangzhou), Institute of Agro-product Safety and Nutrition, Zhejiang Academy of Agricultural Sciences, Hangzhou, China
摘要：
We compared viability, morphology, enzyme activity and gene expression patterns between the chlorine tolerant S. Enteritidis RE 13 and the sensitive strain S. Enteritidis CVCC1806. As expected, viability of strain CVCC1806 was significantly lower than for RE 13 after exposure to 100 mg/L NaClO for 20 min and the sensitive strain displayed severe cell membrane damage and intracellular ATP depletion compared with the tolerant strain. The NAD+/NADH ratios for the sensitive vs tolerant strain were also significantly (p < 0.05) different (3.59 +/- 0.95 vs. 0.88 +/- 0.47) and indicated that tolerance included adaptation to redox perturbations. Transcriptomes of these two strains following chlorine exposure identified significant up-regulation of efflux pump systems (proV/W, OmpF, marA), oxidative stress response (trxC, yhhP, soxS) and DNA repair (dnaQ, polB, yaiV) for the tolerant RE 13 strain. These data indicated that S. Enteritidis RE 13 has a more robust response to oxidative stress that acted to maintain its intracellular redox balance to adapt and survive in the presence of chlorine. Noteworthy, the operon gene cluster of the bacterial microcompartment PDU were up-regulated about 64-fold in tolerant strain RE 13 compared with the sensitive strain, posing a more serious threat to human intestinal health.
摘要：
Background Persistent hyperglycemia decreases the sensitivity of insulin-sensitive organs to insulin, owing to which cells fail to take up and utilize glucose, which exacerbates the progression of type 2 diabetes mellitus (T2DM). lncRNAs' abnormal expression is reported to be associated with the progression of diabetes and plays a significant role in glucose metabolism. Herein, we study the detailed mechanism underlying the functions of lncRNA EPB41L4A-AS1in T2DM. Methods Data from GEO datasets were used to analyze the expression of EPB41L4A-AS1 between insulin resistance or type 2 diabetes patients and the healthy people. Gene expression was evaluated by qRT-PCR and western blotting. Glucose uptake was measured by Glucose Uptake Fluorometric Assay Kit. Glucose tolerance of mice was detected by Intraperitoneal glucose tolerance tests. Cell viability was assessed by CCK-8 assay. The interaction between EPB41L4A-AS1 and GCN5 was explored by RNA immunoprecipitation, RNA pull-down and RNA-FISH combined immunofluorescence. Oxygen consumption rate was tested by Seahorse XF Mito Stress Test. Results EPB41L4A-AS1 was abnormally increased in the liver of patients with T2DM and upregulated in the muscle cells of patients with insulin resistance and in T2DM cell models. The upregulation was associated with increased TP53 expression and reduced glucose uptake. Mechanistically, through interaction with GCN5, EPB41L4A-AS1 regulated histone H3K27 crotonylation in the GLUT4 promoter region and nonhistone PGC1 beta acetylation, which inhibited GLUT4 transcription and suppressed glucose uptake by muscle cells. In contrast, EPB41L4A-AS1 binding to GCN5 enhanced H3K27 and H3K14 acetylation in the TXNIP promoter region, which activated transcription by promoting the recruitment of the transcriptional activator MLXIP. This enhanced GLUT4/2 endocytosis and further suppressed glucose uptake. Conclusion Our study first showed that the EPB41L4A-AS1/GCN5 complex repressed glucose uptake via targeting GLUT4/2 and TXNIP by regulating histone and nonhistone acetylation or crotonylation. Since a weaker glucose uptake ability is one of the major clinical features of T2DM, the inhibition of EPB41L4A-AS1 expression seems to be a potentially effective strategy for drug development in T2DM treatment.
通讯机构：
[Liang Luo] N;National Engineering Research Center for Nanomedicine, College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, China<&wdkj&>Key Laboratory of Molecular Biophysics of the Ministry of Education, College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, China<&wdkj&>Hubei Key Laboratory of Bioinorganic Chemistry and Materia Medica, School of Chemistry and Chemical Engineering, Huazhong University of Science and Technology, Wuhan, China
