Applied Biochemistry and Biotechnology最新文献

{"title":"Biodiesel Production from Soybean Oil Using a Free-Enzyme and Whole-Cell Dual Lipase System as a Biocatalyst.","authors":"Dunchi Xiao, Xun Li, Yu Zhang, Fei Wang","doi":"10.1007/s12010-024-05115-x","DOIUrl":"https://doi.org/10.1007/s12010-024-05115-x","url":null,"abstract":"<p><p>A dual lipase system has been developed to convert soybean oil into biodiesel through synergistic catalysis of Thermomyces lanuginosus lipase (TLL) and Yarrowia lipolytica lipase 2 (YLL) in this study. Pichia pastoris recombinant strains expressing lipases were successfully constructed, and the activities of TLL and YLL in the fermentation supernatant reached 23,142.71 ± 280.54 U/mL and 895.44 ± 27.31 U/mL, respectively. Immediately thereafter, free lipase was used to catalyze the preparation of biodiesel from soybean oil. After optimizing reaction conditions, 80 U/g oil TLL and 20 U/g oil YLL were used to catalyze the production of biodiesel, and a 95.56% biodiesel yield was obtained at 40 °C, 40% moisture content (water/oil, w/w), and stepwise addition of five molar equivalents of methanol. Double lipase plasmids (tll gene and yll gene in different proportions) were constructed in vitro and introduced into P. pastoris to construct a recombinant strain with optimal activity. Under the reaction conditions of 40% moisture content, 8% whole-cell biocatalyst dosage, and stepwise addition of five molar equivalents of methanol, the biodiesel yield reached 95.35% after 24 h at 40 °C. These results show that synergistic catalysis is an effective strategy for biodiesel synthesis and can not only improve the biodiesel yield but also shorten the reaction time. This study provides a scientific basis for biodiesel production by multi-enzyme compounding, with potential industrial applications.</p>","PeriodicalId":465,"journal":{"name":"Applied Biochemistry and Biotechnology","volume":" ","pages":""},"PeriodicalIF":3.1,"publicationDate":"2024-12-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142845464","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A Feasibility Study on 3-D Imaging of Intrahepatic Bile Ducts in Patients with Biliary Atresia Using Airy Beam Excited Two-Photon Microscopy.","authors":"Jiayinaxi Musha, Yilin Zhao, Rong Zhao, Yuan Li, Rongjuan Sun, Pengfei Liu, Jianghua Zhan","doi":"10.1007/s12010-024-05090-3","DOIUrl":"https://doi.org/10.1007/s12010-024-05090-3","url":null,"abstract":"<p><p>The purpose of this study was to utilize a two-photon microscope excitation Airy beam to achieve three-dimensional imaging of intrahepatic bile ducts in BDL mice and patients with biliary atresia (BA). Ten male Balb/c mice aged 6-8 weeks underwent extrahepatic bile duct ligation (BDL), and 10 underwent sham operation as control. After the operation, the mice resulted in symptoms such as jaundice, darkened urine, and weight loss. Taken liver tissues from BDL and control mice and trimmed to 5*5*3 mm³ after 10 days. Sixteen patients with BA were included in this study. Liver transplantation was performed in 12 cases of them; liver hilar and liver margin tissues were taken during the operation. Kasai portoenterostomy (KPE) was performed in 4 cases, and liver margin tissues were taken. Intraoperative liver tissue samples were trimmed to a size of 5*5*5 mm³. The specimens were subjected to tissue fixation, antigen retrieval, antibody incubation, and subsequent tissue hyalinization following the principles of immunofluorescence staining. Subsequently, light-sheet fluorescence microscopy (LSFM) was followed, and intrahepatic bile ducts of the specimen were imaged utilizing Airy beam which was excited with high imaging depth attenuation-compensated two-photon. Deconvolution was applied to image processing to construct a three-dimensional model of intrahepatic bile ducts. Three-dimensional imaging of liver tissue was conducted in both BDL mice and BA patients, and the distribution of intrahepatic bile ducts was visualized. BDL mice exhibited notable widening of intrahepatic bile ducts, accompanied by bile duct hyperplasia. There was no obvious hyperplasia of intrahepatic bile duct in the control group. Significant small bile duct hyperplasia was seen on imaging of the intrahepatic bile ducts in patients with BA. The intrahepatic bile duct was disorganized and hyperplasia especially in patients who performed liver transplantation. The technique of Airy beam three-dimensional reconstruction can effectively image the intrahepatic bile ducts in Balb/c mice and BA patients in three dimensions. This approach contributes to a better understanding of the distribution of intrahepatic bile ducts in BA patients. Moreover, it facilitates the exploration of models that more accurately simulate BA disease by elucidating the distribution of intrahepatic bile ducts in animal models. Understanding the distribution characteristics of intrahepatic bile duct will facilitate the formulation of hilar bile duct microstructure classification, which can guide the operation and evaluate the prognosis better.</p>","PeriodicalId":465,"journal":{"name":"Applied Biochemistry and Biotechnology","volume":" ","pages":""},"PeriodicalIF":3.1,"publicationDate":"2024-12-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142826784","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"EXT1 and Its Methylation Involved in the Progression of Uterine Corpus Endometrial Carcinoma Pathogenesis.","authors":"Hua Chen, Cailing Han, Chunfang Ha","doi":"10.1007/s12010-024-05116-w","DOIUrl":"https://doi.org/10.1007/s12010-024-05116-w","url":null,"abstract":"<p><p>Uterine corpus endometrial carcinoma (UCEC) is one of the most common gynecologic tumors. Due to the high recurrence and metastasis of UCEC, it is crucial for patients to find new biomarkers for diagnosis and therapy. In this study, R software and the TCGA database were used to screen candidate UCEC predictive markers. Western blot and RT-qPCR were performed to detect protein and mRNA expression of EXT1 in UCEC cell lines. In addition, MTT assay, flow cytometry, transwell assay, and wound healing assay were conducted to assess the cell viability, apoptosis, invasion, and migration in UCEC cells. Overlap-extension PCR technique was employed to construct the vector targeting the deletion of the methylated segment of EXT1. The results showed that a total of 11 candidate genes were obtained and EXT1 was identified as a potential target. The expression and methylation levels of EXT1 were both increased in UCEC tissues and cell lines, as well as elevated EXT1 was closely related to the poor prognosis of patients. Besides, the knockdown of EXT1 significantly inhibited the malignant biological behaviors in UCEC cells. Additionally, the current study also found that the deletion of 1559-2146 bp CpG island segment upregulated EXT1 expression and promoted malignant biological behaviors in UCEC cells. Furthermore, the presence of m7G RNA methylation in UCEC cells also was found. In conclusion, the methylation of EXT1 influenced the gene expression, thereby affecting the malignant biological behaviors in UCEC cells and regulating the pathological progression of UCEC.</p>","PeriodicalId":465,"journal":{"name":"Applied Biochemistry and Biotechnology","volume":" ","pages":""},"PeriodicalIF":3.1,"publicationDate":"2024-12-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142823825","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Identification and Validation of Molecular Features of the Anoikis Gene-Related Hub Genes in Nasopharyngeal Carcinoma.","authors":"Dong Li, Lihao Bao, Shaosheng Liu, Ke Ji, Xujiu Xu, Jie Yuan, Guihua Xia","doi":"10.1007/s12010-024-05130-y","DOIUrl":"https://doi.org/10.1007/s12010-024-05130-y","url":null,"abstract":"<p><p>Nasopharyngeal carcinoma (NPC) is a malignant tumor originating from nasopharyngeal mucosa. Anoikis, a form of programmed cell death induced by detachment from the extracellular matrix, normally prevents metastasis. Resistance to anoikis in cancer cells can enhance their metastatic potential. This study identifies anoikis-related genes (ARGs) associated with NPC to elucidate tumorigenesis mechanisms. Analysis of the GSE12452 dataset from GEO revealed 77 differentially expressed ARGs in NPC tissues. GO and KEGG analyses highlighted significant enrichment in apoptosis-related pathways. A PPI network identified MYC, FN1, BRCA1, and FGF2 as Hub genes. Correlation analysis showed MYC positively correlated with activated dendritic cells (p < 0.01) but negatively with naive CD4 T cells (p < 0.001). FN1 was positively correlated with activated dendritic cells (p < 0.01) and negatively with M1 macrophages (p < 0.05). FGF2 negatively correlated with naive CD4 T cells (p < 0.001), while BRCA1 was positively correlated with eosinophils (p < 0.01). GSVA and GSEA indicated that MYC, FN1, BRCA1, and FGF2 were significantly enriched in cell cycle and DNA replication pathways. Immunohistochemistry and qPCR of 50 NPC samples confirmed the overexpression of these genes. Knockdown of MYC, FN1, BRCA1, and FGF2 led to increased tumor cell malignancy, with statistical significance (p < 0.05). This study identifies MYC, FN1, BRCA1, and FGF2 as anoikis-related genes (ARGs) with significant regulatory roles in nasopharyngeal carcinoma (NPC). These ARGs are found to be involved in the development and progression of NPC, suggesting their potential as therapeutic targets for this cancer.</p>","PeriodicalId":465,"journal":{"name":"Applied Biochemistry and Biotechnology","volume":" ","pages":""},"PeriodicalIF":3.1,"publicationDate":"2024-12-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142811769","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"MARCH5 Promotes the Progression of Thyroid cancer by Regulating Mitochondrial Autophagy Protein FUNDC1-mediated Pyroptosis.","authors":"Haili Tang, Jiangang Wang, Guoxiong Ji, Xiaojun Yang, Huan Yang, Xin Chen, Xiaozhou Yang, Huadong Zhao, Xianli He","doi":"10.1007/s12010-024-05113-z","DOIUrl":"https://doi.org/10.1007/s12010-024-05113-z","url":null,"abstract":"<p><p>MARCH5 is a key regulatory factor in mitochondria. However, the expression and function of MARCH5 in thyroid cancer (TC) are not yet clear. The research explores the role and the potential mechanism of MARCH5 in the tumorigenesis of TC. MARCH5 expression were measured by qRT-PCR and Western blot. CCK-8 kits were used to measure the cell viability. Cell scratch assay and Tanswell assay were used to measure cell migration and invasion, respectively. The pyroptosis related proteins (NLRP3, caspase-1, GSDMD) and mitochondrial autophagy related proteins (LC3-II, p62, parkin, pink1) were detected. The mitochondrial ROS GSH, MDA, and SOD were detected using commercial kits. Finally, a TC mouse model was constructed to detect the role of MARCH5 in tumor growth in vivo. The results displayed that the expression of MARCH5 was increased in TC patients and cells, and was significantly correlated with prognosis. Functional studies have found that MARCH5 inhibits oxidative stress levels and mitochondrial autophagy in TPC-1 cells. Further research has found that MARCH5 promotes the progression of thyroid cancer by degrading FUNDC1 and inhibiting the mitochondrial autophagy mediated pyroptosis pathway, regulating cell proliferation, migration, and invasion in TPC-1 cells. More importantly, interference with MARCH5 inhibits tumor growth and further development of TC in vivo. In conclusion, MARCH5 promotes the progression of thyroid cancer by degrading FUNDC1 and inhibiting the mitochondrial autophagy mediated pyroptosis, regulating cell proliferation, migration, and invasion. This study provides new theoretical basis for the treatment and prevention of TC in clinical practice.</p>","PeriodicalId":465,"journal":{"name":"Applied Biochemistry and Biotechnology","volume":" ","pages":""},"PeriodicalIF":3.1,"publicationDate":"2024-12-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142811772","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"MiR-592 Attenuates Tamoxifen Resistance in Breast Cancer Through PIK3CA-Mediated PI3K/AKT/mTOR Signaling Pathway.","authors":"Conghui Jin, Xiangxiang Gao, Jingyi Ni, Baochun Zhang, Zhenxin Wang","doi":"10.1007/s12010-024-05123-x","DOIUrl":"https://doi.org/10.1007/s12010-024-05123-x","url":null,"abstract":"<p><p>Tamoxifen (TAM) is vital in breast cancer (BC) treatment, yet its resistance significantly impairs its efficacy. While miR-592 is known for its suppressive role in BC, its effect on chemotherapy resistance remains unclear. In this study, we observed a significant reduction in miR-592 levels in TAM-resistant BC tissues and cell lines. Low miR-592 expression was significantly associated with advanced TNM stage, lymph node metastasis, and poorer patient survival. Dual-luciferase assay confirmed miR-592 binding to the predicted gene PIK3CA. Increasing miR-592 levels decreased the IC50 of TAM, inhibited cell viability, migration, and invasion, and enhanced apoptosis in vitro, which was mimicked by PIK3CA knockdown and reversed by PIK3CA overexpression. Moreover, miR-592 upregulation suppressed tumor growth and improved TAM responsiveness in vivo. Molecularly, both si-PIK3CA and miR-592 mimics decreased the expression ratios of p-PI3K/PI3K, p-AKT/AKT, and p-mTOR/mTOR, while increasing cleaved caspase-3 and E-cadherin expression in MCF-7/TAM cells. PIK3CA overexpression partially reversed these reductions. In conclusion, our study demonstrates that miR-592 attenuates TAM resistance by inhibiting the PIK3CA-driven PI3K/AKT/mTOR signaling pathway, representing a promising strategy to address chemoresistance in BC.</p>","PeriodicalId":465,"journal":{"name":"Applied Biochemistry and Biotechnology","volume":" ","pages":""},"PeriodicalIF":3.1,"publicationDate":"2024-12-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142805678","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Biodiesel Production from Waste Oil Through Efficient Enzymatic Synthesis Using Yarrowia lipolytica Lipase 2 in the Presence of Glucose, β-Cyclodextrin, or G50.","authors":"Dunchi Xiao, Xun Li, Yu Zhang, Fei Wang","doi":"10.1007/s12010-024-05095-y","DOIUrl":"https://doi.org/10.1007/s12010-024-05095-y","url":null,"abstract":"<p><p>In this study, the liquid lipase Yarrowia lipolytica lipase 2 (YLLip2) expressed by Pichia pastoris GS115 was used to produce biodiesel from waste oil. Four signal peptides were compared to express YLLip2 in P. pastoris, among which SP23 exhibited greater secretion performance. In a 1.3-L bioreactor with FM22 medium for 7 days incubation, the maximum YLLip2 activity and total protein content reached 895.44 ± 27.31 U/mL and 3.83 ± 0.31 g/L, respectively. Under the optimal reaction conditions of 30 °C, 20% moisture content, 50 U/g oil of enzyme dosage, and distributed methanol addition, the reaction yield reached 80.99% after 12 h. To further improve the biodiesel yield, some additives were used to assist with YLLip2. The results showed that adding G50 (approximately 1/20 of YLLip2) increased the yield by approximately 90% after 6 h cultivation without changing the enzyme dosage. Compared with previous studies, the reaction time for biodiesel production from waste oil in this study was significantly shortened. This study provides a workable method for converting low-quality feedstocks containing high free fatty acids into biodiesel using a liquid lipase as the catalyst.</p>","PeriodicalId":465,"journal":{"name":"Applied Biochemistry and Biotechnology","volume":" ","pages":""},"PeriodicalIF":3.1,"publicationDate":"2024-12-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142790675","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Decoding Drug Discovery: Exploring A-to-Z In Silico Methods for Beginners.","authors":"Hezha O Rasul, Dlzar D Ghafour, Bakhtyar K Aziz, Bryar A Hassan, Tarik A Rashid, Arif Kivrak","doi":"10.1007/s12010-024-05110-2","DOIUrl":"https://doi.org/10.1007/s12010-024-05110-2","url":null,"abstract":"<p><p>The drug development process is a critical challenge in the pharmaceutical industry due to its time-consuming nature and the need to discover new drug potentials to address various ailments. The initial step in drug development, drug target identification, often consumes considerable time. While valid, traditional methods such as in vivo and in vitro approaches are limited in their ability to analyze vast amounts of data efficiently, leading to wasteful outcomes. To expedite and streamline drug development, an increasing reliance on computer-aided drug design (CADD) approaches has merged. These sophisticated in silico methods offer a promising avenue for efficiently identifying viable drug candidates, thus providing pharmaceutical firms with significant opportunities to uncover new prospective drug targets. The main goal of this work is to review in silico methods used in the drug development process with a focus on identifying therapeutic targets linked to specific diseases at the genetic or protein level. This article thoroughly discusses A-to-Z in silico techniques, which are essential for identifying the targets of bioactive compounds and their potential therapeutic effects. This review intends to improve drug discovery processes by illuminating the state of these cutting-edge approaches, thereby maximizing the effectiveness and duration of clinical trials for novel drug target investigation.</p>","PeriodicalId":465,"journal":{"name":"Applied Biochemistry and Biotechnology","volume":" ","pages":""},"PeriodicalIF":3.1,"publicationDate":"2024-12-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142765274","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Effect of Sodium Alginate-Bulk Chitosan/Chitosan Nanoparticle Wall Matrix on the Viability of Lactobacillus plantarum Under Simulated Gastrointestinal Fluids.","authors":"Raghda Abdulhussain Kareem, Seyed Hadi Razavi, Zeinab E Mousavi","doi":"10.1007/s12010-024-05105-z","DOIUrl":"10.1007/s12010-024-05105-z","url":null,"abstract":"<p><p>The viability of probiotic cells decreases during passage through the gastrointestinal tract. The process of probiotics encapsulation with sodium alginate and chitosan polymers was carried out to protect the Lactobacillus plantarum in adverse conditions. Lactobacillus plantarum was entrapped in sodium alginate/chitosan (SA/BChi) and sodium alginate/nano-chitosan (SA/NChi) wall materials. Encapsulating L. plantarum with SA/BChi and SA/NChi resulted in a high encapsulation efficiency % of ~ 86.41 to 91.09%. In addition, coating bacteria cells in encapsulants improved the survivability of the cells under the simulated gastrointestinal fluids by ~ 52.61% in SA/Chi and 58.04% in SA/NChi compared to 29% for unencapsulated forms. Probiotic beads under field emission-scanning electron microscopy (FE-SEM) were morphologically compact with a cracked appearance of SA/NChi beads. The Fourier transform-infrared spectroscopy (FT-IR) and differential scanning calorimetry (DSC) showed vigorous electrostatic interaction between polymers, as well as the high melting points, which corroborate the previous investigations in the field for using SA/BChi or SA/NChi as a promising encapsulating agent for ameliorating the survivability of probiotics under harsh conditions. The distinctive properties possessed by the two coatings make them excellent candidates for use as polymeric carriers in probiotic delivery systems.</p>","PeriodicalId":465,"journal":{"name":"Applied Biochemistry and Biotechnology","volume":" ","pages":""},"PeriodicalIF":3.1,"publicationDate":"2024-12-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142765276","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Heterologous Biosynthesis of Taxifolin in Yarrowia lipolytica: Metabolic Engineering and Genome-Scale Metabolic Modeling.","authors":"Yuxin Sui, Yumei Han, Zetian Qiu, Bingyang Yan, Guang-Rong Zhao","doi":"10.1007/s12010-024-05099-8","DOIUrl":"https://doi.org/10.1007/s12010-024-05099-8","url":null,"abstract":"<p><p>Taxifolin, also known as dihydroquercetin (DHQ), is a flavonoid recognized for its potent antioxidant properties and a wide range of biological activities, including anti-tumor, antiviral, and immunomodulatory effects. Conventional extraction and chemical synthesis methods for taxifolin are often limited by low yields and associated environmental concerns. In this study, we investigated the heterologous biosynthesis of taxifolin in Yarrowia lipolytica through a combination of metabolic engineering and genome-scale metabolic modeling (GSM), complemented by flux balance analysis (FBA). We engineered Yarrowia lipolytica by introducing key biosynthetic genes and successfully synthesized taxifolin using naringenin (NAR) as a substrate, chosen for its low cost. Fermentation experiments demonstrated an optimal taxifolin yield of 10% at a substrate concentration of 200 mg/L naringenin, with a maximum yield of 26.4 mg/L taxifolin at 1 g/L naringenin. To further enhance production, we applied a marker-free Cre-loxP-based gene integration method, allowing stable genomic integration of key genes, which increased taxifolin yield to 34.9 mg/L at 1 g/L naringenin. Additionally, intermediate metabolites eriodictyol (ERI) and dihydrokaempferol (DHK) accumulated to concentrations of 89.2 mg/L and 21.7 mg/L, respectively. Furthermore, we integrated metabolic data into a GSM and applied FBA to optimize the taxifolin biosynthetic pathway. Through Pareto frontier analysis, sensitivity analysis, flux variability analysis, and single gene deletion simulations, we identified key genetic modifications that significantly enhanced taxifolin yield. Overexpression of GND1 and IDP2 increased yields by 94% and 155%, respectively, while knockout of LIP2 led to a 46% increase. Using tri-baffled shake flasks to improve oxygen supply resulted in a 120% yield increase, whereas YPG medium decreased yield by 59%, validating our model's accuracy. To ensure stable and efficient gene expression, we integrated multi-copy constructs into the ribosomal DNA (rDNA) locus of Yarrowia lipolytica, doubling taxifolin production. These results demonstrate the effectiveness of GSM and FBA in addressing bottlenecks in microbial taxifolin biosynthesis and provide a basis for future optimization and large-scale production.</p>","PeriodicalId":465,"journal":{"name":"Applied Biochemistry and Biotechnology","volume":" ","pages":""},"PeriodicalIF":3.1,"publicationDate":"2024-12-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142765247","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}