Applied Biochemistry and Biotechnology最新文献

Background: YTH domain-containing protein 1 (YTHDC1) has been confirmed to be involved in regulating esophageal cancer (EC) progression. However, whether YTHDC1 mediates the radioresistance of EC and its potential molecular mechanism still need to be further explored.

Methods: The mRNA and protein levels of YTHDC1, ubiquitin-specific peptidase 10 (USP10) and polo-like kinase 1 (PLK1) were determined by qRT-PCR and western blot. Cell proliferation, invasion, migration, radiosensitivity, and apoptosis were detected by CCK8 assay, transwell assay, colony formation assay, and flow cytometry. The level of autophagy-related marker LC3B was examined using western blot. The interaction between USP10 and YTHDC1 or PLK1 was assessed by RIP assay, Co-IP assay and ubiquitination assay. Animal experiments were performed to explore the role of YTHDC1 in vivo.

Results: YTHDC1 was increased expression in EC tissues and cells. Silencing of YTHDC1 suppressed EC cell proliferation, migration and invasion, while promoted apoptosis, radiosensitivity and autophagy. YTHDC1 could stabilize USP10 mRNA level, and USP10 increased PLK1 expression by deubiquitination. Further analysis showed that PLK1 overexpression reversed the regulation of YTHDC1 knockdown on EC cell proliferation, invasion, radiosensitivity and autophagy. Besides, YTHDC1 knockdown could inhibit EC tumorigenesis and improve radiosensitivity in vivo via inactivating USP10/PLK1 axis.

Conclusion: Targeted inhibition of YTHDC1/USP10/PLK1 axis may be an effective measure to inhibit EC progression and improve radiosensitivity.

Scorpions have long been utilized in traditional medicine for their therapeutic properties, including anti-cancer, analgesic, anti-thrombotic, and immune-modulating properties. This study aimed to optimize extraction conditions, partial purification, structural characterization and biological activity of glycoproteins from the scorpion (Buthus martensii Karsch) body. Under the optimized conditions, the extraction yield of scorpion glycoproteins (SGP's) reached 12.57 ± 0.7%. Subsequently, two novel glycoproteins, SGP-1a and SGP-1b, were isolated through DEAE-52 anion exchange chromatography and Sephadex G-75 gel filtration, with average molecular weights of 46.8 kDa and 10.3 kDa, respectively. Compositional analysis revealed SGP-1a and SGP-1b were rich in monosaccharides (Glu, Gal, Rha) and amino acids (Ser, Lys, Ala, Leu, Arg, Phe), and were characterized as homogeneous acidic protein-bound heteropolysaccharides. FTIR spectrum confirmed that SGP-1a and SGP-1b presence of glycated proteins, β-elimination reaction elucidated that SGP's contained N-glycosidic linkage. According to the CD analysis, SGP-1a contains 7.4% α-helix, 34.1% β-sheet, 13.2% β-turn, 34.6% random coil and SGP-1b contains 4.7% α-helix, 46.8% β-strand, 47.2% random coil. Bioactivity assays demonstrated significant anti-inflammatory activity against COX-2 (IC50: 21.14 ± 1.73 µg/mL for SGP-1a; 18.58 ± 1.54 µg/mL for SGP-1b) Meanwhile, both the SGPs exhibit strong antioxidant activity, the IC₅₀ of SGP-1a and SGP-1b against DPPH, ABTS, and hydroxyl free radicals were 1.75 mg/mL, and 0.79 mg/mL, 1.56 mg/mL, and 0.79 mg/mL, 0.53 mg/mL and 1.69 mg/mL respectively. These findings unveil the molecular basis of BmK's medicinal properties, providing novel leads for anti-inflammatory drug development.

Abtract: The filamentous cyanobacterium Limnothrix sp. strain SK1-2-1 is capable of producing pentadecane (a C15 alkane) and phycocyanin. The aim of this study was to develop a method for the production and recovery of extracellular substances (skECS), containing a novel acidic extracellular polysaccharide (skEPS) as a biopolymer, and to analyze the properties and functions of the obtained skECS. Analyses showed that skECS had a high molecular weight of 1.46 × 10⁶ g/mol and comprised 28.2% (w/w) sugar and 30.2% (w/w) protein. By contrast, skEPS contained glucose (38.4 mol%), mannose (35.3 mol%), and rhamnose (14.8 mol%). skECS was characterized using native PAGE and atomic force microscopy. Aqueous skECS solutions at concentrations ranging from 0.25 to 1.00 mg mL^-1 showed antioxidant capacity. Additionally, blue-light irradiation of SK1-2-1 cells enhanced skEPS production and promoted simultaneous cell flocculation. The accumulation of candidate gene transcripts involved in skEPS biosynthesis in response to blue- and white-light irradiation showed that the expression of sesA (sk0924), which encodes the blue-light sensor substance cyclic-di-GMP, was induced by blue light. Furthermore, nine genes related to the Wzy-dependent pathway, which were inferred to act in an sesA-dependent manner, were non-uniformly induced under blue-light irradiation, with varying degrees of expression. These results reveal novel properties and functions of skECS, light-specific induction of production, and their potential as useful materials for biorefineries.

Sucrose isomerase (SIM) faces challenges in industrial applications due to its limited stability and reusability. To address these issues, a copper ion-mediated hybrid nanoflower immobilization system was developed for a rationally engineered SIM mutant (Q310E). Under optimized synthesis conditions (50 h, 120 mM Cu²⁺, SIM-to-Cu²⁺ ratio of 0.012 mg/mM, 4℃), the resulting SIM@Cu-NFs exhibited a well-defined hierarchical structure and high batch-to-batch reproducibility. The immobilized enzyme showed significantly enhanced stability, retaining over 80% activity after 1 h at pH 4.0-8.0, over 70% activity after 3 h at 50 ℃, and over 70% activity after 45 days of storage at 4℃. Moreover, it maintained 55.9% activity after 6 reuse cycles and approximately 45% after 12 cycles. Kinetic analysis revealed a 38.5% increase in catalytic efficiency (K_cat/K_m), along with improved substrate affinity (K_m decreased from 53.5 to 45.0 ± 0.7 mM) and a higher reaction rate (V_max increased from 1609 to 1950 ± 28 µM·min^- 1). These enhancements are attributed to the stabilizing effect of Cu²⁺ coordination bonds and the favorable microenvironments within the nanoflower architecture. The SIM@Cu-NF system demonstrates high operational stability, reusability, and catalytic efficiency, showing promising potential for industrial applications in enzyme-based biocatalysis.

Non-small cell lung cancer (NSCLC) remains a leading cause of cancer-related deaths worldwide, necessitating the identification of novel therapeutic targets. Mesoderm posterior bHLH transcription factor 1 (MESP1) has been implicated in various developmental processes, but its role in cancer, particularly NSCLC, is poorly understood. Given the emerging evidence linking MESP1 to cellular processes relevant to cancer biology, investigating its regulatory mechanisms in NSCLC could provide critical insights for developing new therapies. This study employed quantitative real-time PCR (qRT-PCR) to assess the mRNA levels of MESP1, ubiquitin specific peptidase 7 (USP7), and clusters of differentiation 163 (CD163). Western blotting was used to analyze the protein expression of MESP1 and USP7. Cellular proliferation was evaluated through colony-forming assays, while apoptosis was quantified using flow cytometry. Mitochondrial membrane potential was measured by JC-1 staining, and reactive oxygen species (ROS) levels were also analyzed via flow cytometry. Additionally, colorimetric assays were utilized to determine malondialdehyde (MDA), total iron, and Fe²⁺ levels. The in vivo effects of MESP1 silencing on NSCLC progression were examined using a xenograft mouse model. GST-pull down assay, Co-immunoprecipitation (Co-IP) assay, and ubiquitination assay were conducted to explore the interaction between USP7 and MESP1. The expression of both MESP1 and USP7 was found to be upregulated in NSCLC tissues and cells when compared with normal lung tissues and normal human bronchial epithelial cells. Knockdown of MESP1 significantly inhibited NSCLC cell proliferation, induced apoptosis and promoted features associated with ferroptosis. Moreover, MESP1 silencing suppressed M2 macrophage polarization and tumor formation. Mechanistically, USP7 was identified to stabilize MESP1 protein expression through its deubiquitinating activity. Overexpression of MESP1 attenuated the inhibitory effects of USP7 silencing on NSCLC cell proliferation and M2 macrophage polarization and also mitigated the promoting effects of USP7 knockdown on apoptosis and the induction of features associated with ferroptosis. USP7 stabilized MESP1 to promote the malignant progression of NSCLC. The findings highlight the potential of targeting the USP7-MESP1 axis as a novel therapeutic strategy for NSCLC.

Keliu Pill (KLW) is a Chinese medicine formula that has been shown to be effective in treating non-small cell lung cancer (NSCLC) patients. However, the potential molecular mechanism remains unclear. To explore the underlying mechanism of Keliu Pill (KLW) in treating NSCLC, the network pharmacology was applied to explore the potential mechanism of KLW in the treatment of lung cancer. With oral bioavailability (OB) ≥ 30% and drug-like index (DL) ≥ 0.18 as the filter criteria, the compounds of 6 traditional Chinese medicines (TCMs) from KLW were retrieved on Traditional Chinese Medicine Systematic Pharmacology Database and Analysis Platform (TCMSP) and supplemented by Traditional Chinese Medicine Comprehensive Database (TCMID). The potential targets corresponding to the active components of the TCMs mentioned above were obtained from TCMSP. Subsequently, the potential underlying action mechanisms of KLW on NSCLC predicted by the network pharmacology analyses were experimentally validated in Lewis lung cancer cell inoculation mice models and co-culture models with Lewis lung cancer cell and tumor-associated macrophages (TAMs). After database search and screening, a total of 20 active ingredients of Astragalus Membranaceus, 21 of Sophare Tonkinensis Radix et Rhizoma, 1 of Mylabris, 3 of Eupolyphaga, 1 of Leech, and 3 of Gecko were obtained. After deleting duplicates, 49 active ingredients were obtained. Thirty important active ingredients in KLW were sorted out by referring to relevant literature. The KEGG pathway enrichment analysis based on the DAVID platform showed that the VEGF-C signaling pathway may be the core signaling pathway associated with KLW in the treatment of NSCLC. The network pharmacological analysis demonstrated that the various components of KLW acted on NSCLC through 29 potential targets such as VEGF-C, VEGF-D, MAPK8, MAPK1, and AKT1. The experiments in vivo indicated KLW could inhibit the transcription and translation of VEGF-C and VEGF-D in tumor tissues. KLW could reduce the proportion of activated TAMs in the tissue. KLW-contained serum may decrease the expression level of VEGF-C and VEGF-D and inhibit the lymphatic lumen formation of human lymphatic endothelial cells. VEGFC and VEGFD were confirmed as the potential KLW-associated targets for NSCLC. KLW may inhibit lymphatic angiogenesis of lung cancer by regulating the function of TAMs.

In recent years, bacterial quorum quenching (QQ) has emerged as an effective strategy for reducing biofouling in membrane bioreactors (MBRs). Understanding microbial community dynamics is crucial for developing effective QQ strategies, as changes in these communities can significantly influence the risk of biofouling in the sludge. This study systematically investigates the formation, mechanisms, and regulatory strategies related to biofouling in MBRs. It offers a comprehensive analysis of quorum sensing (QS) mechanisms within microbial communities and their biofouling tendencies. Moreover, the interactions between quorum sensing, extracellular polymerization, and membrane biofouling are discussed. Additionally, the short-term addition of exogenous QQ was found to temporarily cause a reduction in the sludge's QQ capabilities, thereby increasing its susceptibility to membrane biofouling. The study concludes with future perspectives on managing biofouling in membrane bioreactors and provides recommendations for further research on leveraging QS-MBR systems to mitigate membrane biofouling.