摘要：
Addition of chemical structural information in enzymatic reactions has proven to be significant for accurate enzyme function prediction. However, such chemical data lack systematic feature mining and hardly exist in enzyme-related databases. Therefore, global mining of enzymatic reactions will offer a unique landscape for researchers to understand the basic functional mechanisms of natural bioprocesses and facilitate enzyme function annotation. Here, we established a new knowledge base called EnzyMine, through which we propose to elucidate enzymatic reaction features and then link them with sequence and structural annotations. EnzyMine represents an advanced database that extends enzyme knowledge by incorporating reaction chemical feature strategies, strengthening the connectivity between enzyme and metabolic reactions. Therefore, it has the potential to reveal many new metabolic pathways involved with given enzymes, as well as expand enzyme function annotation. Database URL: http://www.rxnfinder.org/enzymine/.
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.
作者机构：
[He, Lei; Yu, Ning; Chen, Ying] Chinese Academy of Inspection and Quarantine, Beijing, 100176, PR China;School of Food and Health, Beijing Technology and Business University, Beijing, 100048, PR China;Graduate School of Food Science and Engineering, Jilin Agricultural University, Beijing, 130118, PR China;College of Life Science and Technology, Wuhan Polytechnic University, Wuhan, 430023, PR China;[Hu, Qian] Chinese Academy of Inspection and Quarantine, Beijing, 100176, PR China<&wdkj&>School of Food and Health, Beijing Technology and Business University, Beijing, 100048, PR China
通讯机构：
[Ying Chen] C;Chinese Academy of Inspection and Quarantine, Beijing, 100176, PR China
摘要：
目的·探究miR-185-5p在分枝杆菌感染的巨噬细胞中对自噬的调控作用和对胞内分枝杆菌生存的影响。方法·使用卡介苗(Bacillus Calmette-Guérin,BCG)感染人单核细胞...展开更多 目的·探究miR-185-5p在分枝杆菌感染的巨噬细胞中对自噬的调控作用和对胞内分枝杆菌生存的影响。方法·使用卡介苗(Bacillus Calmette-Guérin,BCG)感染人单核细胞白血病细胞(THP-1)诱导分化的巨噬细胞,通过超速离心法分离细胞培养上清液中的外泌体;通过Western blotting、纳米颗粒跟踪分析(nanoparticle tracking analysis,NTA)、透射电子显微镜(transmission electron microscopy,TEM)分别对外泌体的表面标志蛋白、大小和形态进行鉴定,通过实时荧光定量PCR(quantitative polymerase chain reaction,qPCR)检测BCG感染的巨噬细胞内及外泌体中miR-185-5p的表达水平。外泌体使用PKH26染料进行荧光染色并与巨噬细胞共培养,观察巨噬细胞对外泌体的吞噬情况。通过模拟物(mimic)和抑制剂(inhibitor)实现miR-185-5p在巨噬细胞内的过表达和抑制作用,通过菌落形成单位(colony-forming unit,CFU)实验验证miR-185-5p对巨噬细胞内BCG生长存活的影响。通过Western blotting检测自噬标志物微管相关蛋白1轻链3(microtubule-associated protein 1 light chain 3,LC3)蛋白的表达,研究过表达、抑制miR-185-5p以及使用自噬抑制剂氯喹(chloroquine,CQ)对巨噬细胞自噬的影响;进一步使用SensGFP-StubRFP-LC3自噬双荧光慢病毒检测miR-185-5p对自噬流的影响。使用TargetScan、miRDB和Starbase数据库对miR-185-5p的靶基因进行预测,并使用注释、可视化和综合发现数据库(Database for Annotation,Visualization and Integrated Discovery,DAVID)工具进行基因本体(Gene Ontology,GO)分析和京都基因与基因组百科全书(Kyoto Encyclopedia of Genes and Genomes,KEGG)功能富集分析。结果·超速离心法成功分离出巨噬细胞外泌体,Western blotting成功检测到外泌体标志蛋白白细胞分化抗原9(cluster differentiation 9,CD9)、肿瘤易感基因101(tumor susceptibility gene 101,TSG101);通过TEM、NTA鉴定,外泌体为双层膜结构,直径约150 nm。BCG感染后,miR-185-5p在巨噬细胞内和外泌体中的表达均显著上调(P=0.000),且其在BCG感染的巨噬细胞内的表达水平呈时间依赖性升高。外泌体与巨噬细胞共培养,可被巨噬细胞吞噬。与mimic和inhibitor的各自对照相比,mimic显著上调miR-185-5p的表达(P=0.000),而inhibitor抑制miR-185-5p的表达(P=0.002)。使用mimic引起巨噬细胞内BCG的CFU数量显著增加(P=0.000),而inhibitor则使CFU数量减少(P=0.041)。BCG感染使巨噬细胞中LC3Ⅱ蛋白的表达上调,而mimic可降低LC3Ⅱ的表达,inhibitor可升高LC3Ⅱ的表达。在CQ作用下,使用inhibitor抑制miR-185-5p仍能显著增强巨噬细胞中LC3Ⅱ的表达。自噬流检测结果显示,相比于各自的对照组,mimic组自噬体的生成明显减少(P=0.034),而inhibitor组自噬体的生成明显增加(P=0.042)。通过靶基因预测以及GO和KEGG功能富集分析发现,miR-185-5p可能通过靶向核受体亚家族1组D成员1(nuclear receptor subfamily 1 group D member 1,NR1D1)、Ras相关蛋白Rab-35(Ras-related protein Rab-35,RAB35)、细胞分裂控制蛋白42同源物(cell division control protein 42 homolog,CDC42)等基因发挥自噬调节作用。结论·分枝杆菌感染诱导巨噬细胞内及外泌体中miR-185-5p表达上调,miR-185-5p通过抑制自噬体的生成抑制巨噬细胞的自噬过程,从而促进细胞内分枝杆菌的生长及存活。收起
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.
作者机构：
[骆健美; 黄蓉; 白杰; 张鹤渐; 李从雨] College of Biotechnology, Tianjin University of Science & Technology, Tianjin 300457, China;[刘晓楠; 黄蓉; 江会锋; 王千; 白杰; 张鹤渐; 李从雨; 张磊] Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China;[张磊] College of Life Science and Technology, Wuhan Polytechnic University, Wuhan 430023, Hubei, China
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.
