Search Results (41)

Curcumin, the major bioactive polyphenol derived from the edible rhizome turmeric (Curcuma longa L.), is recognized for its health-promoting properties. Despite well-documented antioxidant effects, its molecular mechanisms, particularly those involving post-transcriptional regulation, remain incompletely understood. This in vitro study identifies a novel microRNA-mediated pathway contributing to the antioxidant activity of curcumin in human hepatic LO2 cells. Curcumin treatment downregulated the stress-responsive microRNA miR-22-3p. Bioinformatics analysis and a dual-luciferase reporter assay identified malonyl-CoA-acyl carrier protein transacylase (MCAT), a mitochondrial enzyme, as a direct target of miR-22-3p. Modulation of this axis reduced intracellular reactive oxygen species (ROS), enhanced total reducing capacity, increased activities of key antioxidant enzymes (SOD, CAT, GPx), and improved mitochondrial bioenergetics without altering membrane potential. Crucially, siRNA-mediated knockdown of MCAT attenuated the ROS-scavenging effect of curcumin. These findings reveal a mechanistic pathway wherein curcumin downregulates miR-22-3p, resulting in upregulation of MCAT and enhanced mitochondrial antioxidant defense. This work broadens the understanding of curcumin’s bioactivity from direct radical scavenging to include the post-transcriptional fine-tuning of mitochondrial metabolism. The study establishes a molecular framework for further exploration of curcumin’s potential in alleviating oxidative stress. Full article

Malonyl-CoA decarboxylase (MCD) is an enzyme that controls malonyl-CoA levels and regulates fatty acid synthesis and oxidation. Although its physiological relevance in peripheral tissues is well known, the role of MCD in the central nervous system remains poorly understood. MCD is expressed in mitochondria, cytosol, and peroxisomes and may be regulated by PPAR-α, AMPK, and SIRT4 in tissues such as muscle, liver and kidney. In the brain, MCD expression varies during development and can respond to nutritional states. Inherited MCD deficiency (malonic aciduria) leads to the toxic accumulation of malonic acid and predominantly affects the central nervous system. The underlying mechanisms leading to brain damage in MCD patients remain unclear. Conversely, pharmacological modulation of MCD activity has been studied in obesity, diabetes, and ischemic injury, highlighting its therapeutic potential. There are still major gaps regarding MCD cellular distribution, regulatory pathways, and metabolic interaction with CPT1c (carnitine palmitoyltransferase 1c) in neural metabolism. A deeper understanding of the role of MCD in brain physiology and pathology may indicate novel therapeutic strategies targeting metabolic disorders that involve altered malonyl-CoA dynamics. Here, we discuss the current knowns and unknowns regarding MCD physiology, regulation, and pathophysiology, emphasizing brain aspects. Full article

The evolution of type I polyketide synthase (T1PKS) assembly lines remains poorly understood. Through systematic mining of polyene biosynthetic gene clusters, we identified a novel eurocidin biosynthetic pathway capable of producing identical compounds with divergent loading module architectures, thereby capturing an evolutionary transitional state. Biochemical analysis revealed unprecedented functional reprogramming of acyltransferase (AT) domains, shifting substrate specificity from extender units (malonyl-CoA) to starter units (acyl-CoA). This paradigm shift enables direct initiation of polyketide chain assembly via AT-mediated loading of starter units, thereby elucidating the origin of extant AT-initiated assembly lines and establishing AT functional plasticity as a novel mechanism for polyketide structural diversification. Parallel evolution of ketosynthase (KS) domains through KSS→KSQ mutations further diversified initiation strategies. Applying this evolutionary insight, we engineered the candicidin pathway by replacing its native aromatic-starting bimodule with a starter-selective monomodule from eurocidin, generating aliphatic-starting analogs. This demonstrates that evolution-inspired AT reprogramming provides a rational framework for modifying polyketide starter units, expanding structural diversity, and enhancing therapeutic potential. Full article

Background/Objectives: Paeonia delavayi, a high-altitude-adapted medicinal and oil-producing plant, exhibits broad elevational distribution. Understanding how environmental factors regulate its growth across altitudes is critical for optimizing cultivation and exploiting its economic potential. Methods: In this study, we conducted a comprehensive Iso-Seq and RNA-seq analysis to elucidate the transcriptional profile across diverse altitudes and three seed developmental stages. Results: Using Pacbio full-length cDNA sequencing, we identified 39,267 full-length transcripts, with 80.03% (31,426) achieving successful annotation. RNA-seq analysis uncovered 11,423 and 9565 differentially expressed genes (DEGs) in response to different altitude and developmental stages, respectively. KEGG analysis indicated that pathways linked to fatty acid metabolism were notably enriched during developmental stages. In contrast, pathways associated with amino acid and protein metabolism were significantly enriched under different altitudes. Furthermore, we identified 34 DEGs related to fatty acid biosynthesis, including genes encoding pivotal enzymes like biotin carboxylase, carboxyl transferase subunit alpha, malonyl-CoA-acyl carrier protein transacylase, 3-oxoacyl-ACP reductase, 3-hydroxyacyl-ACP dehydratase, and stearoyl-ACP desaturase enoyl-ACP reductase. Additionally, ten DEGs were pinpointed as potentially involved in high-altitude stress response. Conclusions: These findings provide insights into the molecular mechanisms of fatty acid biosynthesis and adaptation to high-altitude stress in peony seeds, providing a theoretical foundation for future breeding programs aimed at enhancing seed quality. Full article

The introduction of heterologous pathways into microbial cell compartments offers several potential advantages, including increasing enzyme concentrations and reducing competition with native pathways, making this approach attractive for producing complex metabolites like fatty acids and fatty alcohols. However, measuring subcellular concentrations of these metabolites remains technically challenging. Here, we explored 3-hydroxypropionic acid (3-HP), readily quantifiable and sharing the same precursors—acetyl-CoA, NADPH, and ATP—with the above-mentioned products, as a reporter metabolite for peroxisomal engineering in the yeast Komagataella phaffii. To this end, the malonyl-CoA reductase pathway for 3-HP production was targeted into the peroxisome of K. phaffii using the PTS1-tagging system, and further tested with different carbon sources. Thereafter, we used compartmentalized 3-HP production as a reporter system to showcase the impact of different strategies aimed at enhancing the peroxisomal NADPH pool. Co-overexpression of genes encoding a NADPH-dependent redox shuttle from Saccharomyces cerevisiae (IDP2/IDP3) significantly increased 3-HP yields across all substrates, whereas peroxisomal targeting of the S. cerevisiae NADH kinase Pos5 failed to improve 3-HP production. This study highlights the potential of using peroxisomal 3-HP production as a biosensor for evaluating peroxisomal acetyl-CoA and NAPDH availability by simply quantifying 3-HP, demonstrating its potential for peroxisome-based metabolic engineering in yeast. Full article

Bibenzyl compounds are one of the most important bioactive components of natural medicine. However, Dendrobium officinale as a traditional herbal medicine is rich in bibenzyl compounds and performs functions such as acting as an antioxidant, inhibiting cancer cell growth, and assisting in neuro-protection. The biosynthesis of bibenzyl products is regulated by bibenzyl synthase (BBS). In this study, we have cloned the cDNA gene of the bibenzyl synthase (DoBS1) from D. officinale using PCR with degenerate primers, and we have identified a novel type III polyketide synthase (PKS) gene by phylogenetic analyses. In a series of perfect experiments, DoBS1 was expressed in Escherichia coli, purified and some catalytic properties of the recombinant protein were investigated. The molecular weight of the recombinant protein was verified to be approximately 42.7 kDa. An enzyme activity analysis indicated that the recombinant DoBS1-HisTag protein was capable of using 4-coumaryol-CoA and 3 malonyl-CoA as substrates for dihydroresveratrol (DHR) in vitro. The Vmax and Km of the recombinant protein for DHR were 3.57 ± 0.23 nmol·min⁻¹·mg⁻¹ and 0.30 ± 0.08 mmol, respectively. The present study provides further insights into the catalytic mechanism of the active site in the biosynthetic pathway for the catalytic production of dihydroresveratrol by bibenzylase in D. officinale. The results can be used to optimize a novel biosynthetic pathway for the industrial synthesis of DHR. Full article

The marine Streptomyces harbor numerous biosynthetic gene clusters (BGCs) with exploitable potential. However, many secondary metabolites cannot be produced under laboratory conditions. Co-culture strategies of marine microorganisms have yielded novel natural products with diverse biological activities. In this study, we explored the metabolic profiles of co-cultures involving Streptomyces sp. 2-85 and Cladosporium sp. 3-22—derived from marine sponges. Combining Global Natural Products Social (GNPS) Molecular Networking analysis with natural product database mining, 35 potential antimicrobial metabolites annotated were detected, 19 of which were exclusive to the co-culture, with a significant increase in production. Notably, the Streptomyces-Fungus interaction led to the increased production of borrelidin and the discovery of several analogs via molecular networking. In this study, borrelidin was first applied to combat Saprolegnia parasitica, which caused saprolegniosis in aquaculture. We noted its superior inhibitory effects on mycelial growth with an EC₅₀ of 0.004 mg/mL and on spore germination with an EC₅₀ of 0.005 mg/mL compared to the commercial fungicide, preliminarily identifying threonyl-tRNA synthetase as its target. Further analysis of the associated gene clusters revealed an incomplete synthesis pathway with missing malonyl-CoA units for condensation within this strain, hinting at the presence of potential compensatory pathways. In conclusion, our findings shed light on the metabolic changes of marine Streptomyces and fungi in co-culture, propose the potential of borrelidin in the control of aquatic diseases, and present new prospects for antifungal applications. Full article

Resveratrol, a phenylpropanoid compound, exhibits diverse pharmacological properties, making it a valuable candidate for health and disease management. However, the demand for resveratrol exceeds the capacity of plant extraction methods, necessitating alternative production strategies. Microbial synthesis offers several advantages over plant-based approaches and presents a promising alternative. Yarrowia lipolytica stands out among microbial hosts due to its safe nature, abundant acetyl-CoA and malonyl-CoA availability, and robust pentose phosphate pathway. This study aimed to engineer Y. lipolytica for resveratrol production. The resveratrol biosynthetic pathway was integrated into Y. lipolytica by adding genes encoding tyrosine ammonia lyase from Rhodotorula glutinis, 4-coumarate CoA ligase from Nicotiana tabacum, and stilbene synthase from Vitis vinifera. This resulted in the production of 14.3 mg/L resveratrol. A combination of endogenous and exogenous malonyl-CoA biosynthetic modules was introduced to enhance malonyl-CoA availability. This included genes encoding acetyl-CoA carboxylase 2 from Arabidopsis thaliana, malonyl-CoA synthase, and a malonate transporter protein from Bradyrhizobium diazoefficiens. These strategies increased resveratrol production to 51.8 mg/L. The further optimization of fermentation conditions and the utilization of sucrose as an effective carbon source in YP media enhanced the resveratrol concentration to 141 mg/L in flask fermentation. By combining these strategies, we achieved a titer of 400 mg/L resveratrol in a controlled fed-batch bioreactor. These findings demonstrate the efficacy of Y. lipolytica as a platform for the de novo production of resveratrol and highlight the importance of metabolic engineering, enhancing malonyl-CoA availability, and media optimization for improved resveratrol production. Full article

Isoflavones are predominantly found in legumes and play roles in plant defense and prevention of estrogen-related diseases. Genistein is an important isoflavone backbone with various biological activities. In this paper, we describe how a cell factory that can de novo synthesize genistein was constructed in Saccharomyces cerevisiae. Different combinations of isoflavone synthase, cytochrome P450 reductase, and 2-hydroxyisoflavone dehydratase were tested, followed by pathway multicopy integration, to stably de novo synthesize genistein. The catalytic activity of isoflavone synthase was enhanced by heme supply and an increased intracellular NADPH/NADP⁺ ratio. Redistribution of the malonyl-CoA flow and balance of metabolic fluxes were achieved by adjusting the fatty acid synthesis pathway, yielding 23.33 mg/L genistein. Finally, isoflavone glycosyltransferases were introduced into S. cerevisiae, and the optimized strain produced 15.80 mg/L of genistin or 10.03 mg/L of genistein-8-C-glucoside. This is the first de novo synthesis of genistein-8-C-glucoside in S. cerevisiae, which is advantageous for the green industrial production of isoflavone compounds. Full article

Soybean [Glycine max (L.) Merr.] isoflavones, which are secondary metabolites with various functions, are included in food, cosmetics, and medicine. However, the molecular mechanisms regulating the glycosylation and malonylation of isoflavone glycoconjugates remain unclear. In this study, we conducted an RNA-seq analysis to compare soybean genotypes with different isoflavone contents, including Danbaek and Hwanggeum (low-isoflavone cultivars) as well as DB-088 (high-isoflavone mutant). The transcriptome analysis yielded over 278 million clean reads, representing 39,156 transcripts. The analysis of differentially expressed genes (DEGs) detected 2654 up-regulated and 1805 down-regulated genes between the low- and high-isoflavone genotypes. The putative functions of these 4459 DEGs were annotated on the basis of GO and KEGG pathway enrichment analyses. These DEGs were further analyzed to compare the expression patterns of the genes involved in the biosynthesis of secondary metabolites and the genes encoding transcription factors. The examination of the relative expression levels of 70 isoflavone biosynthetic genes revealed the HID, IFS, UGT, and MAT expression levels were significantly up/down-regulated depending on the genotype and seed developmental stage. These expression patterns were confirmed by quantitative real-time PCR. Moreover, a gene co-expression analysis detected potential protein–protein interactions, suggestive of common functions. The study findings provide valuable insights into the structural genes responsible for isoflavone biosynthesis and accumulation in soybean seeds. Full article

Stevia rebaudiana (Bertoni) is a highly valuable crop for the steviol glycoside content in its leaves, which are no-calorie sweeteners hundreds of times more potent than sucrose. The presence of health-promoting phenolic compounds, particularly flavonoids, in the leaf of S. rebaudiana adds further nutritional value to this crop. Although all these secondary metabolites are highly desirable in S. rebaudiana leaves, the genes regulating the biosynthesis of phenolic compounds and the shared gene network between the regulation of biosynthesis of steviol glycosides and phenolic compounds still need to be investigated in this species. To identify putative candidate genes involved in the synergistic regulation of steviol glycosides and phenolic compounds, four genotypes with different contents of these compounds were selected for a pairwise comparison RNA-seq analysis, yielding 1136 differentially expressed genes. Genes that highly correlate with both steviol glycosides and phenolic compound accumulation in the four genotypes of S. rebaudiana were identified using the weighted gene co-expression network analysis. The presence of UDP-glycosyltransferases 76G1, 76H1, 85C1, and 91A1, and several genes associated with the phenylpropanoid pathway, including peroxidase, caffeoyl-CoA O-methyltransferase, and malonyl-coenzyme A:anthocyanin 3-O-glucoside-6″-O-malonyltransferase, along with 21 transcription factors like SCL3, WRK11, and MYB111, implied an extensive and synergistic regulatory network involved in enhancing the production of such compounds in S. rebaudiana leaves. In conclusion, this work identified a variety of putative candidate genes involved in the biosynthesis and regulation of particular steviol glycosides and phenolic compounds that will be useful in gene editing strategies for increasing and steering the production of such compounds in S. rebaudiana as well as in other species. Full article

Methylmalonic aciduria and homocystinuria type C protein (MMACHC) is required by the body to metabolize cobalamin (Cbl). Due to its complex structure and cofactor forms, Cbl passes through an extensive series of absorptive and processing steps before being delivered to mitochondrial methyl malonyl-CoA mutase and cytosolic methionine synthase. Depending on the cofactor attached, MMACHC performs either flavin-dependent reductive decyanation or glutathione (GSH)-dependent dealkylation. The alkyl groups of Cbl have to be removed in the presence of GSH to produce intermediates that can later be converted into active cofactor forms. Pathogenic mutations in the GSH binding site, such as R161Q, R161G, R206P, R206W, and R206Q, have been reported to cause Cbl diseases. The impact of these variations on MMACHC’s structure and how it affects GSH and Cbl binding at the molecular level is poorly understood. To better understand the molecular basis of this interaction, mutant structures involving the MMACHC-MeCbl-GSH complex were generated using in silico site-directed point mutations and explored using molecular dynamics (MD) simulations. The results revealed that mutations in the key arginine residues disrupt GSH binding by breaking the interactions and reducing the free energy of binding of GSH. Specifically, variations at position 206 appeared to produce weaker GSH binding. The lowered binding affinity for GSH in the variant structures could impact metabolic pathways involving Cbl and its trafficking. Full article

Numerous investigations have shown that insoluble dietary fiber (IDF) has a potentially positive effect on obesity due to a high-fat diet (HFD). Our previous findings based on proteomic data revealed that high-purity IDF from soybean residue (okara) (HPSIDF) prevented obesity by regulating hepatic fatty acid synthesis and degradation pathways, while its intervention mechanism is uncharted. Consequently, the goal of this work is to find out the potential regulatory mechanisms of HPSIDF on hepatic fatty acid oxidation by determining changes in fatty acid oxidation-related enzymes in mitochondria and peroxisomes, the production of oxidation intermediates and final products, the composition and content of fatty acids, and the expression levels of fatty acid oxidation-related proteins in mice fed with HFD. We found that supplementation with HPSIDF significantly ameliorated body weight gain, fat accumulation, dyslipidemia, and hepatic steatosis caused by HFD. Importantly, HPSIDF intervention promotes medium- and long-chain fatty acid oxidation in hepatic mitochondria by improving the contents of acyl-coenzyme A oxidase 1 (ACOX1), malonyl coenzyme A (Malonyl CoA), acetyl coenzyme A synthase (ACS), acetyl coenzyme A carboxylase (ACC), and carnitine palmitoyl transferase-1 (CPT-1). Moreover, HPSIDF effectively regulated the expression levels of proteins involved with hepatic fatty acid β-oxidation. Our study indicated that HPSIDF treatment prevents obesity by promoting hepatic mitochondrial fatty acid oxidation. Full article

3-Hydroxypropionic acid (3-HP) is an important intermediate compound in the chemical industry. Green and environmentally friendly microbial synthesis methods are becoming increasingly popular in a range of industries. Compared to other chassis cells, Yarrowia lipolytica possesses advantages, such as high tolerance to organic acid and a sufficient precursor required to synthesize 3-HP. In this study, gene manipulations, including the overexpression of genes MCR-N${}_{Ca}$, MCR-C${}_{Ca}$, GAPN${}_{Sm}$, ACC1 and ACS${}_{Se}^{L641P}$ and knocking out bypass genes MLS1 and CIT2, leading to the glyoxylate cycle, were performed to construct a recombinant strain. Based on this, the degradation pathway of 3-HP in Y. lipolytica was discovered, and relevant genes MMSDH and HPDH were knocked out. To our knowledge, this study is the first to produce 3-HP in Y. lipolytica. The yield of 3-HP in recombinant strain Po1f-NC-14 in shake flask fermentation reached 1.128 g·L⁻¹, and the yield in fed-batch fermentation reached 16.23 g·L⁻¹. These results are highly competitive compared to other yeast chassis cells. This study creates the foundation for the production of 3-HP in Y. lipolytica and also provides a reference for further research in the future. Full article

Lysine malonylation (Kmal) is an evolutionarily conserved post-translational modification (PTM) that has been demonstrated to be involved in cellular and organismal metabolism. However, the role that Kmal plays in response to drought stress of the terrestrial cyanobacteria N. flagelliforme is still unknown. In this study, we performed the first proteomic analysis of Kmal in N. flagelliforme under different drought stresses using LC-MS/MS. In total, 421 malonylated lysine residues were found in 236 different proteins. GO and KEGG enrichment analysis indicated that these malonylated proteins were highly enriched in several metabolic pathways, including carbon metabolism and photosynthesis. Decreased malonylation levels were found to hinder the reception and transmission of light energy and CO₂ fixation, which led to a decrease in photosynthetic activity. Kmal was also shown to inhibit the flux of the TCA cycle and activate the gluconeogenesis pathway in response to drought stress. Furthermore, malonylated antioxidant enzymes and antioxidants were synergistically involved in reactive oxygen species (ROS) scavenging. Malonylation was involved in lipid degradation and amino acid biosynthesis as part of drought stress adaptation. This work represents the first comprehensive investigation of the role of malonylation in dehydrated N. flagelliforme, providing an important resource for understanding the drought tolerance mechanism of this organism. Full article