Search Results (104)

Soil hypoxia caused by waterlogging severely restricts kiwifruit growth, and screening waterlogging-tolerant rootstocks and analyzing their mechanisms are of great significance for industrial development. In this study, waterlogging-tolerant Actinidia valvata ‘Shuixiu’ was used as the test material and Actinidia chinensis ‘Hongyang’ as the control. Waterlogging stress was simulated artificially, and physiological measurements combined with transcriptome sequencing were used to explore its waterlogging tolerance regulatory characteristics based on respiratory metabolism. The results showed that the waterlogging tolerance of ‘Shuixiu’ was significantly better than that of ‘Hongyang’. It upregulated sucrose synthase and α/β-amylase genes and inhibited the continuous up-regulation of trehalose-6-phosphate synthase genes, leading to significant accumulation of glucose-6-phosphate, a key glycolytic substrate. Some members of glycolytic key gene families, such as glucose-6-phosphate isomerase and phosphofructokinase, were upregulated in ‘Shuixiu’, which increased phosphoglycerate kinase activity and accumulated 3-phosphoglyceric acid and pyruvate, ensuring efficient conversion of carbon sources to ATP. Some members of core tricarboxylic acid cycle gene families, such as pyruvate dehydrogenase and citrate synthase, were upregulated in ‘Shuixiu’, with significantly higher pyruvate dehydrogenase activity and acetyl coenzyme A content, maintaining partial aerobic respiration capacity. Some members of the alanine transaminase gene family were upregulated in ‘Shuixiu’ to enhance alanine fermentation, resulting in a significant reduction in root ethanol accumulation. This study clarified the core respiratory metabolic regulatory characteristics of kiwifruit in response to waterlogging and provided key targets and a theoretical basis for molecular breeding of waterlogging-tolerant rootstocks. Full article

Erythroid acute myeloid leukemia (AML) cell line OCI-M2 expresses a particular oncogenic network: IRF6, in concert with ETV2 and HEY1, aberrantly activates NKL homeobox gene NKX2-4, which in turn represses megakaryocytic lineage factor FLI1. Interestingly, in keratinocytes, IRF6 is able to bind glucose which promotes IRF6-dimerization and thus alters its binding site selection. Here, we used OCI-M2 as a model to investigate the role of glucose level and IRF6 in leukemogenesis. Treatment of OCI-M2 with high glucose or 2-deoxy-glucose resulted in the downregulation of IRF6 and NKX2-4, and the upregulation of FLI1, indicating that glucose-mediated dimerization of IRF6 altered its reported autoactivation. The screening of this cell line for genes encoding glycolytic enzymes identified aberrant overexpression of glucose-6-phosphate isomerase (GPI) and phosphofructokinase L (PFKL), which were targeted by genomic amplification and chromothripsis-like alterations, respectively. Furthermore, GPI was activated by NKX2-4 and ETV2, and PFKL by ETV2. Finally, siRNA-mediated downregulation of PFKL resulted in elevated glucose levels, suppressed expression of IRF6 and NKX2-4, and activated FLI1. Thus, we connected an oncogenic regulatory network with deregulated glycolytic enzymes and glucose metabolism, thereby establishing a new in vitro model to develop novel therapeutic avenues in AML subsets. Full article

D-allulose is a rare low-calorie sugar with considerable health benefits and industrial potential. Compared with chemical synthesis and free enzyme catalysis, microbial production using engineered cells offers a low-cost and highly stable solution. Therefore, we investigated the reaction pathway underlying the synthesis of D-allulose from D-glucose. Specifically, the enhancement of glucose isomerase-catalyzed reactions and their role in D-allulose synthesis were evaluated. First, a mutant strain with significantly increased glucose isomerase from Anoxybacillus kamchatkensis G10 (AGGI) expression was obtained through ultraviolet mutagenesis combined with high-throughput flow cytometry. A 4.55-fold increase in AGGI activity and a D-fructose conversion yield of 51.2% were obtained. A dual-enzyme pathway was subsequently constructed by co-expressing AGGI and D-allulose 3-epimerase (DAEase) in the optimized host. After balancing the catalytic requirements of both enzymes through optimization of reaction conditions, CRISPR-associated transposase was employed to efficiently integrate the glucose transporter gene galP into the genome, further enhancing substrate supply. The final engineered Escherichia coli strain achieved a D-allulose conversion rate of 15% from 20 g/L D-glucose. This demonstrates the crucial role of glucose isomerase in microbial D-allulose production and advances the optimization and development of D-allulose synthesis strategies using D-glucose as a substrate. Full article

Serial femtosecond crystallography (SFX) using X-ray free electron lasers (XFELs) enables the determination of room-temperature structures of biological macromolecules without radiation damage. The accuracy of detector geometry parameters, including the crystal-to-detector distance (CTDD), is critical for reliable data processing. In SFX experiments, the CTDD may shift during data collection due to changes in the experimental setup or installation of the sample delivery system. Such CTDD variations can affect the quality of SFX datasets; however, their impact has not been fully elucidated in the context of SFX data processing. In this study, we investigated the influence of CTDD variations on SFX datasets collected at Pohang Accelerator Laboratory X-ray Free Electron Laser (PAL-XFEL) with thermolysin, lysozyme, and glucose isomerase crystals processed by four indexing algorithms. At the optimized CTDD, the distribution of unit cell parameters exhibited a Gaussian pattern; however, it became distorted as the CTDD deviated further from the optimal value. Data analysis indicated that the CTDD tolerance for successful data processing and structure determination was approximately ±3–5 mm from the optimized CTDD. These findings provide insight into indexing behavior in SFX data processing at PAL-XFEL and offer practical guidance for improving data processing efficiency. Full article

Fructooligosaccharides (FOS) represent a major source of prebiotic compounds. They are widely used in functional foods for their ability to modify intestinal microbiota in animals and humans. To address the significant issue of fructooligosaccharide production being influenced by glucose concentration, this study designed a dual-enzymatic co-catalysis system for glucose isomerase (GI) and a mutant FTase (FTase^142P-242K). This system successfully increased the FOS synthesis rate (42.31 to 55.51%, w/w). Glucose isomerase catalyzes the isomerization of glucose to fructose, and the subsequent release of fructose from the active site permits the enzyme to re-enter its catalytic cycle. The optimal conditions for catalysis were found at 45 °C, pH 5.5, and 1 mM Ba²⁺. In contrast, the optimal fermentation process was established at 25 °C and induction with 1 mM IPTG. Finally, the efficient production of FOS using low-cost byproduct molasses was achieved. Fermentation optimization of the dual-enzyme system resulted in FOS yield of 53.92% (w/w), a significant increase (44.54%, w/w) from the yield obtained using single-enzyme catalysis. Based on the research, a novel and sustainable approach for high-yield synthesis of Fructooligosaccharides involves minimizing the inhibitory effect of glucose produced during sucrose transformation. Full article

Arthritis in K/BxN mice is provoked by pathogenic autoantibodies to glucose-6-phosphate isomerase (G6PI), which is a ubiquitously expressed enzyme that is present in cells, in the circulation and on articular cartilage. When G6PI autoantibodies (auto-Abs) deposit on the articular cartilage of K/BxN mice, arthritis ensues due to the activation of various components of the innate immune system. Recent studies have investigated the in vivo efficacy of recombinant fragment-crystallizable (Fc) protein-based therapeutics. Many recombinant Fc proteins evaluated provide protection against inflammation in mouse models of arthritis, such as the K/BxN serum-transfer model. More recently, rFc-µTP-L309C, a recombinant human IgG1-Fc with an additional point mutation at position L309C fused to the human IgM tailpiece to form a hexamer, has been shown to ameliorate the arthritis in K/BxN mice. Additional studies have shown that rFc-µTP-L309C has multiple effects that work together to ameliorate the arthritis, including inhibition of neutrophil migration into the joint, inhibition of IL-1β production, downregulation of Th1 and Th17 cells, and increases in T regulatory cells and synovial fluid IL-10. In this work, rFc-µTP-L309C was shown to effectively prevent arthritis in the K/BxN serum-transfer model, significantly downregulate inflammatory cytokines/chemokines, and ameliorate the arthritis in the endogenous K/BxN model. This amelioration of the arthritis was associated with a significant decrease in autoantibody levels, which was independent of the neonatal Fc receptor (FcRn). rFc-µTP-L309C was shown to specifically inhibit G6PI autoantibody secretion from B-cells with a concomitant increase in TGFβ and decrease in B-cell activating factor (BAFF). These new findings suggest that rFc-µTP-L309C may provide a therapeutic benefit for other antibody-mediated autoimmune diseases through its effects on B-cells. Full article

Disorders in soybean seed-filling can lead to wrinkled seeds, affecting yield and quality. Previous studies have demonstrated that some soybean cultivars from Jiamusi, Heilongjiang Province (cold-temperate continental monsoon, ~3.5 °C mean annual temperature, ~530 mm precipitation) exhibit seed-filling disorders when cultivated in Shenyang, Liaoning Province (mid-temperate semi-humid continental monsoon, ~8.3 °C, ~610 mm). However, the causes and regulatory mechanisms remain unclear. In this study, Henong 76 (a soybean cultivar with seeds less prone to wrinkling) and Heihe 43 (a soybean cultivar with seeds prone to wrinkling) were used as experimental materials. They were sown simultaneously in Jiamusi and Shenyang, respectively, to explore the causes of seed-filling disorders in Heihe 43. The results indicated that there were significant differences in the contents of soluble sugars and starch, as well as in the activities of sucrose synthase and invertase, between the seeds of Henong 76 and Heihe 43 grown in Shenyang. However, no significant differences were found between them in Jiamusi. Transcriptomic and metabolomic analyses suggested that genes related to controlling starch hydrolysis (isoamylase, α-amylase, and glycogen phosphorylase) and sucrose synthesis and decomposition (sucrose synthase, invertase, glucose-6-phosphate isomerase, and phosphoglucomutase) in Heihe 43 were upregulated in Shenyang. In contrast, genes regulating plant hormone signal transduction (auxin, gibberellin, abscisic acid, and cytokinin) were generally downregulated. These changes led to differences in metabolites, resulting in the occurrence of seed-filling disorders. Furthermore, we analyzed the climatic conditions of the two cultivars during the soybean seed-filling period. The results indicated that high temperature might be the primary meteorological factor contributing to the occurrence of seed-filling disorders. All results indicated that the insufficient accumulation of sugars in seeds due to exposure to high temperatures during the seed-filling period is the primary cause of the prone-to-wrinkling phenomenon of the Heihe 43 cultivar under the ecological conditions of Shenyang. Full article

This work reviews the complex role of the enzyme triosephosphate isomerase (TIM) (EC 5.3.1.1) within the context of diabetes, a prevalent metabolic disorder. It summarizes the main biochemical pathways, cellular mechanisms, and molecular interactions that highlight both the function of TIM and its implications in diabetes pathophysiology, particularly focusing on its regulatory role in glucose metabolism and insulin secretion. TIM’s involvement is detailed from its enzymatic action in glycolysis, influencing the equilibrium between dihydroxyacetone phosphate and glyceraldehyde-3-phosphate, to its broader implications in cellular metabolic processes. The article highlights how mutations in TIM can lead to metabolic inefficiencies that exacerbate diabetic conditions. It discusses the interaction of TIM with various cellular pathways, including its role in the ATP-sensitive potassium channels in pancreatic beta cells, which are crucial for insulin release. Moreover, we indicate the impact of oxidative stress in diabetes, noting how TIM is affected by reactive oxygen species, which can disrupt normal cellular functions and insulin signaling. The enzyme’s function is also tied to broader cellular and systemic processes, such as membrane fluidity and cellular signaling pathways, including the mammalian target of rapamycin, which are critical in the pathogenesis of diabetes and its complications. This review emphasizes the dual role of TIM in normal physiological and pathological states, suggesting that targeting TIM-related pathways could offer novel therapeutic strategies for managing diabetes. It encourages an integrated approach to understanding and treating diabetes, considering the multifaceted roles of biochemical players such as TIM that bridge metabolic, oxidative, and regulatory functions within the body. Full article

Hexokinase catalyzes the first rate-limiting step glycolysis. However, the roles of hexokinase 2 (HK2) in asthma remain incompletely understood. This study aimed to investigate metabolic alterations in asthma, focusing on the expression, function and regulation of HK2. In this study, non-targeted metabolomics analysis of 20 asthma patients and 15 healthy controls identified metabolic alterations in asthma, particularly in the glycolytic pathways. Consistently, HK2 expression was elevated in both asthma individuals and mice with allergic airway inflammation. Airway epithelium–specific HK2 knockdown and pharmacological inhibition with 2-deoxy-D-glucose (2-DG) significantly attenuated airway inflammation and hyperresponsiveness in mice induced by ovalbumin/ lipopolysaccharide. Mechanistic analyses demonstrated that HK2 regulates epithelial apoptosis and inflammation via interaction with peptidylprolyl isomerase F (PPIF), independent of voltage-dependent anion channel 1 (VDAC1). Asthma is associated with metabolic reprogramming, characterized by alterations in lipid and glucose metabolism. These findings establish HK2 plays a crucial role in asthma pathogenesis by promoting airway epithelial apoptosis and inflammation in asthma, suggesting its potential as a therapeutic target. Full article

The occurrence of anthocyanins in rice (Oryza sativa) and barley (Hordeum vulgare) varies among cultivars, with pigmented varieties (e.g., black rice and purple barley) accumulating higher concentrations due to genetic and environmental factors. The biosynthesis of anthocyanins is regulated by a complex network of structural and regulatory genes. Key enzymes in the pathway include chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), dihydroflavonol 4-reductase (DFR), anthocyanidin synthase (ANS), and UDP-glucose flavonoid 3-O-glucosyltransferase (UFGT). These genes are tightly controlled by transcription factors (TFs) from the MYB, bHLH (basic helix–loop–helix), and WD40 repeat families, which form the MBW (MYB-bHLH-WD40) regulatory complex. In rice, OsMYB transcription factors such as OsMYB3, OsC1, and OsPL (Purple Leaf) interact with OsbHLH partners (e.g., OsB1, OsB2) to activate anthocyanin biosynthesis. Similarly, in barley, HvMYB genes (e.g., HvMYB10) coordinate with HvbHLH TFs to regulate pigment accumulation. Environmental cues, such as light, temperature, and nutrient availability, further modulate these TFs, influencing the production of anthocyanin. Understanding the genetic and molecular mechanisms behind the biosynthesis of anthocyanins in rice and barley provides opportunities for the development of biofortification strategies that enhance their nutritional value. Full article

As global milk production continues to rise, the disposal of expired milk contributes to environmental pollution and valuable resource wastage. This study presents the development of a novel L-arabinose isomerase, designated BmAIase12, and its application in the enzymatic recycling of expired milk. BmAIase12 exhibited a specific activity of 10.7 U/mg and showed optimal performance at 50 °C and pH 7.0. Furthermore, it exhibited higher activity than most other L-arabinose isomerases. It converted D-galactose into D-tagatose with a high conversion ratio of 53.3% after 48 h at 50 °C. The conversion efficiency of expired milk to D-tagatose was recorded at 40.62%, resulting in a maximum tagatose yield of 1.625 g/L. This was accomplished through the incorporation of β-galactosidase (120 U/mL) and Saccharomyces cerevisiae (30 mg/mL) to hydrolyze lactose and metabolize glucose, followed by the addition of 3 U/mL of BmAIase12. Ultimately, following purification, the purity of tagatose was determined to be 98%, and the final yield was 29.8%. These results suggest that BmAIase12 may serve as a promising enzyme for D-tagatose production due to its high conversion yield. Full article

Phosphoglucose isomerase (PGI), which catalyzes the interconversion of glucose 6-phosphate (G6P) and fructose 6-phosphate (F6P), plays a key role in regulating carbohydrate synthesis and catabolism in plant growth and development. In higher plants, two PGI isoenzymes, plastidic (PGIp) and cytosolic (PGIc), have diverged significantly in sequence, structure, activity, and functional roles, despite catalyzing the same reaction. However, whether they experience distinct selection pressures during intraspecific population differentiation remains unknown. In this study, we identified the PGIc and PGIp genes in castor beans, an important industrial and horticultural crop, and revealed their different expression patterns across tissues, particularly during seed development. Population genetic analyses (Tajima’s D, ML-HKA, and CODEML) detected strong evidence of positive selection on RcPGIc, but not RcPGIp during domestication. Four positively selected sites in RcPGIc (114T, 310T, 338A, and 613S) were inferred with posterior probabilities > 95% in BEB analysis, and two of them (114T and 613S) were found to be significantly associated with higher seed oil content, suggesting that these two sites could potentially influence oil accumulation in castor seeds. This study provides the first evidence of differential selection pressures on PGIc and PGIp during intraspecific population differentiation, offering new insights into their functional divergence. Full article

Erythritol has been widely used in the food industry, which predominantly synthesizes it via microbial fermentation, in which Yarrowia lipolytica serves as the preferred candidate chassis strain. However, the wild-type strain of Y. lipolytica exhibits several limitations, including suboptimal industrial performance and elevated levels of by-products, which pose significant challenges in biomanufacturing processes. It is significant to understand the synthesis mechanism of erythritol for improving the capacity of erythritol production by Y. lipolytica. In this study, a mutant exhibiting high erythritol production and stable genetic performance was obtained via a combination of UV and atmospheric and room-temperature plasma mutagenesis. Some key genes related to erythritol production were identified through comparative transcriptome analysis of the mutant strain, revealing significant changes in their expression levels. Individual overexpression of the genes encoding ribose-5-phosphate isomerase, glucose-6-phosphate-1-epimerase, adenylate kinase, and alcohol dehydrogenase in Y. lipolytica Po1g enhanced erythritol production, demonstrating the critical role of each gene in erythritol production. This finding elucidates the molecular mechanism underlying the improved erythritol yield in the mutant strain. The Y. lipolytica mutant C1 produced 194.47 g/L erythritol in a 10 L fermenter with a productivity of 1.68 g/L/h during batch fermentation, surpassing the wild-type strain and reducing the cultivation time by 21 h. It is significant to understand the mechanism of erythritol synthesis for improving erythritol production and its application in industrial-scale production. Full article

D-tagatose is a functional sweetener with glucose-regulating and prebiotic properties, but its bioproduction from D-galactose faces many limitations, particularly the high production costs. In particular, the current biosynthesis of D-tagatose suffers from thermal instability and the substrate selectivity issues of L-arabinose isomerase (L-AI) required to convert D-galactose into D-tagatose. In this study, recombinant Escherichia coli BW25113/pQE-80L-araA_F118M/F279I expressing double mutant L-AI was immobilized to enhance its stability and reusability. The optimal conditions for whole-cell catalysis were 60 °C, pH 6.5, 5 mM Mn²⁺, and 20 h, with a yield of 55.2 g/L of D-tagatose. Immobilization with 3% sodium alginate and 2% CaCl₂ retained 90% of the production efficiency displayed by free cells. Notably, the immobilized cells exhibited enhanced heat resistance (60–70 °C) and operational stability, retaining 76% activity after five cycles. The D-tagatose production was further increased to 129.43 g/L by increasing the substrate concentration to 250 g/L. Compared to free cells, immobilized cells retained 83.6% of the initial yield up to 10 batches. This study presents a cost-effective and sustainable method for the production of D-tagatose using optimized whole-cell catalysis through immobilization, which paves the way to solve industrial challenges such as thermal instability and low substrate efficiency. Full article

Serial crystallography (SX) enables the determination of biologically relevant structures at room temperature while minimizing radiation damage. During SX experiments, the beam center on diffraction images can shift due to X-ray beam movements or detector displacement. Consequently, the geometry file for the beam center is optimized; however, the effects of deviations from the optimal position on data processing efficiency remain unclear. This study examines how changes in the beam center influence data quality by analyzing the indexing efficiency and structure refinement of lysozyme and glucose isomerase datasets, considering shifts in the beam center parameter. The results revealed that as the beam center deviated farther from its optimal position, the indexing efficiency declined, with the extent of the effect varying significantly across indexing algorithms. XDS and MOSFLM algorithms maintained high indexing efficiencies (>90%) for shifts of ≤4 pixels (688 μm) and ≤2 pixels (344 μm), respectively, compared to data processed at the optimized beam center. Conversely, the DirAx and XGANDALF algorithms exhibited indexing efficiencies below 90% for a two-pixel shift in the beam center. These findings enhance our understanding of how beam center shifts affect SX data processing and provide valuable insights for developing effective data processing strategies. Full article