Search Results (3,476)

Aging is one of the most complex biological processes, which leads to a gradual decline in the function of organs, tissues and cells, and significant increases in the risks of many age-associated diseases, including cancer, neurodegenerative disorders, and cardiovascular diseases. Protein biomarkers have attracted increasing attention in research on aging and age-related diseases. Considering the fact that proteins are large heterogenous biomolecules due to coding polymorphisms, alternative RNA splicing and post-translational modifications (PTMs), including glycosylation, phosphorylation, and methylation, mass spectrometry (MS)-based top-down proteomics (TDP) is a powerful technology that allows for measuring proteins without proteolysis, thus characterizing intact forms of proteins, which provides information on primary sequences, including their modifications. This review provides an overview of TDP technologies, with a particular focus on the separation, ionization, and fragmentation of intact proteins and introduces the most recent applications of TDP to the discovery of proteoform-resolved biomarkers associated with aging and age-related diseases. Full article

Deciphering the coloration mechanism of natural-colored cocoons in the domestic silkworm (Bombyx mori) is of great importance for the green and sustainable development of the sericulture industry. In this study, 14 silkworm strains were selected for studying differences in the coloration mechanism of diverse cocoon colors, and the present research carried out integrated investigations from three aspects: pigment content, differences in gene expression levels, and gene structural variation. The results demonstrated that pigment accumulation presented distinct tissue-specific and strain-specific characteristics. The middle silk gland (MSG) acts as the primary locus for pigment deposition: silkworm strains forming yellow or red cocoons accumulate carotenoids at high levels in this tissue, whereas those producing green cocoons show abundant flavonoid enrichment here. Analysis of gene expression profiles indicated that the expression patterns of core transporter genes are highly correlated with the spatial distribution of pigments. The expression level of CBP gene in the MSG is over 10-fold higher than that in the midgut (MG) among yellow/red cocoon strains. The pivotal glycosylation gene UGT86 displayed remarkably elevated expression in the MSG relative to other tissues across all green cocoon silkworm strains. The CBP gene acts as a core regulatory factor governing the transport of carotenoid pigments, and notable disparities existed in the coding region of the gene among silkworm strains with different cocoon colors. In contrast to yellow and red cocoon strains, the transcription start site of CBP gene is displaced in silkworm varieties that form green or white cocoons. In summary, this study clarified the expression patterns and variations in key pigment deposition-related genes at the population level for the first time and provided data references for the study of the biological basis and coloration mechanism of diverse cocoon colors. Full article

Glycosylation is one of the most prevalent post-translational modifications of membrane proteins and plays a central role in regulating their structure and function. Unlike many existing reviews that address glycosylation in a system-wide context, this review focuses specifically on membrane proteins and examines how glycosylation shapes their functional behavior and clinical relevance. Because membrane proteins are exposed to the extracellular environment, glycans on their surface directly influence protein folding, receptor organization, and interactions with ligands and immune components. These diverse effects can be understood within a common mechanistic framework in which glycosylation modulates protein conformation, receptor clustering, and membrane organization, thereby altering signaling, adhesion, transport, and immune recognition. We discuss how N-linked and O-linked glycosylation regulate major classes of membrane proteins across these processes. Particular attention is given to disease-associated alterations in glycosylation, especially in cancer, immune and inflammatory disorders, and metabolic disease. For instance, glycosylation-dependent stabilization of PD-L1 and modulation of receptor signaling, such as EGFR, illustrate how glycan modifications contribute to immune evasion and therapeutic response. We further consider the clinical implications of membrane protein glycosylation, including its roles in biomarker development and as a potential target for therapeutic intervention. Advances in glycoproteomic technologies have enabled increasingly detailed characterization of site-specific glycosylation, although significant analytical challenges remain, particularly for membrane proteins. Overall, this review highlights membrane protein glycosylation as a dynamic regulatory layer that links molecular mechanisms to functional outcomes and clinical applications. Full article

Background/Objective: Immunoglobulin G (IgG) N-glycosylation is an important regulator of immune function and systemic inflammation and has been associated with cardiometabolic diseases. However, little is known about how IgG glycosylation changes during the course of acute coronary syndrome (ACS) and whether these alterations relate to lipid-lowering response after the initiation of statin therapy. The primary aim of this study was to investigate IgG N-glycosylation following ACS and evaluate its association with response to atorvastatin therapy defined as baseline LDL cholesterol reduction of ≥50%. Methods: In this prospective cohort study, 79 statin-naïve patients hospitalized for the first episode of ACS and treated with atorvastatin 80 mg daily after percutaneous coronary intervention were followed longitudinally. Plasma samples were collected at admission (acute phase), discharge (subacute phase), and follow-up (chronic phase). A control group of 21 individuals received atorvastatin for primary prevention. IgG was isolated from plasma, and N-glycans were released, fluorescently labeled with 2-aminobenzamide, and analyzed using hydrophilic interaction-based ultra-high-performance liquid chromatography with fluorescence detection. Derived glycan traits were calculated, including agalactosylated (G0), monogalactosylated (G1), digalactosylated (G2), core fucosylated (F), bisected (B), and sialylated (S) glycans. Results: No significant differences in derived IgG glycan traits were observed between ACS patients and controls at baseline or follow-up. Within the ACS group, a longitudinal analysis revealed significant increases in G0 and F and a decrease in G2 between the acute and chronic phases. A total of 65% of patients achieved ≥50% reduction in LDL cholesterol (LDL-C), whereas only 22% reached the guideline-recommended LDL-C target of <1.4 mmol/L. Patients achieving ≥50% LDL-C reduction exhibited consistently higher G0 and lower G2 and S across disease phases. In a subgroup of patients with baseline LDL-C >3.9 mmol/L, those who failed to achieve ≥50% LDL-C reduction had significantly lower G0 and higher S across all time points. Conclusions: Specific glycan traits are associated with the degree of LDL-C reduction achieved during statin therapy, particularly in patients with high baseline LDL-C. These findings suggest that IgG glycosylation patterns may reflect biological phenotypes associated with differential lipid-lowering responsiveness after ACS. Full article

Ischemic stroke induces complex molecular responses that disrupt subcellular organelles’ function and contribute to brain injury, yet the temporal changes of organelle-specific transcriptomic remodeling remain to be investigated. In this study, we performed in silico analysis of publicly available transcriptomic data from isolated brain microvessels of transient middle cerebral artery occlusion (tMCAO) mouse model. Using in silico approaches, we analyzed differential gene expression at 24 h (acute phase) and 7 d (intermediate phase) post-stroke, focusing on mitochondria, endoplasmic reticulum (ER), and Golgi apparatus. Functional enrichment (Gene Ontology, KEGG) and protein–protein interaction network analyses were performed. Our analysis of the data revealed that at 24 h post-stroke, all three organelles exhibited marked transcriptional remodeling, where mitochondrial pathways showed disrupted metabolic and redox regulation; ER pathways indicated activation of biosynthetic processes, stress signaling, and ferroptosis; and Golgi-related genes reflected altered vesicular trafficking and glycosylation. By 7 d, mitochondrial alterations subsided, whereas ER and Golgi pathways displayed downregulation of metabolic and neuronal signaling processes, indicating persistent dysfunction and incomplete microvascular recovery. Phase-specific drug–gene interaction analysis will be useful to understand temporal organelle-associated transcriptional organization and to guide future investigations of neurovascular remodeling after ischemic stroke. Full article

Background/Objectives: Onion (Allium cepa) peems are an underutilized by-product rich in polyphenols. This study evaluated the physicochemical profile, and bioactive potential (antidiabetic, antimicrobial, antioxidant, and anticoagulant) of Moroccan red onion peels using integrated in vivo, in vitro, and in silico approaches. Methods: Moisture, pH, ash content, and mineral elements were determined, followed by phytochemical screening and three extractions: decoction E0, aqueous Soxhlet E1, and hydroethanolic Soxhlet E2 (70/30; ethanol/water, v/v). The measurement of polyphenols, flavonoids, and tannins was carried out using colorimetric methods, while the molecular profile was studied by high-performance liquid chromatography coupled to ultraviolet detection and electrospray ionization mass spectrometry (HPLC/UV-ESI-MS). Biological activities were determined using 2,2-diphenyl-1-picrylhydrazyl, ferric reducing antioxidant power, and total antioxidant capacity assays (in vitro antioxidant); microdilution (antimicrobial); prothrombin time and activated partial thromboplastin time (anticoagulant); and α-amylase/α-glucosidase enzymatic inhibition and oral glucose tolerance tests on normoglycemic rats. Also, acute toxicity was evaluated, and molecular interactions between these proteins and ligands (docking, molecular dynamics, and MM-PBSA) were analyzed. Results: Physicochemical analyses showed an acidic pH (3.06) and high ash content (15.21%), with the concentration of regulated elements remaining within FAO/WHO limits. The extractive content was between 6.90% E0 and 19.18% E2. The E1 extract had the maximum amount of total polyphenols (178.95 mg GAE/g); on the other hand, E2 was the richest in flavonoids by 121.43 mg QE/g. The HPLC/ESI-MS analysis of E0 revealed 20 compounds, among which flavonoids (84.93%) were predominant, with isorhamnetin (30.26%), followed by quercetin and its glycosylated forms. E1 showed the most potent antioxidant effects (IC₅₀ DPPH, 22.38 µg/mL, as that of ascorbic acid). The antibacterial activity of E0 was especially potent towards Enterobacter cloacae and Pseudomonas aeruginosa (MIC 75 µg/mL). A mild dose-dependent anticoagulant effect was seen. Antidiabetic activity was found to be outstanding: α-amylase (IC₅₀ 62.75 µg/mL) and α-glucosidase (IC₅₀ 8.49 µg/mL, stronger than acarbose) inhibitions were corroborated in vivo by a considerable decrease in the glycemic area under the curve. The molecular docking study in silico demonstrated strong molecular interactions, especially for quercetin 4′-O-glucoside with good binding energies. Conclusions: A. cepa peels from Morocco can be considered a safe plant matrix containing bioactive flavonoids with strong antioxidant and selective antimicrobial activities and promising antidiabetic effects, supported by molecular modeling. Full article

Leaves of Actinidia chinensis Planch. var. deliciosa (A.Chev.) A.Chev. (A. deliciosa) represent agro-industrial byproducts with potential for valorization. The present study evaluated the metabolomics profiling, cytotoxicity, genotoxicity, and antigenotoxicity of the methanolic extract of A. deliciosa leaves. The metabolomics profiling was determined using an untargeted metabolomic approach employing UPLC-HRMS. Cytotoxicity, genotoxicity, and antigenotoxicity were assessed in Chinese hamster ovary K1 (CHO-K1) cells using the in vitro cytokinesis-block micronucleus (CBMN) assay. The metabolic profile of A. deliciosa leaf extracts revealed the presence of three major classes of secondary/specialized metabolites: proanthocyanidins, flavonols, and triterpenoid saponins. Medium-polar metabolites were monomeric fla-van-3-ols, such as (+)-catechin and (−)-epicatechin, oligomeric procyanidins and prodelphinidins, and flavonols. Certain glycosylated flavonols and their derivatives, such as myricetin, quercetin, and kaempferol. Low-polarity metabolites were characterized by low-polarity triterpenoids such as maslinic, corosolic, oleanolic, and ursolic acids. At concentrations of 37.5, 75, and 150 µg/mL, the extract did not significantly increase micronuclei frequency compared to untreated control cells, indicating an absence of genotoxic potential. Moreover, co-treatment of CHO-K1 cells with the extract and mitomycin C (MMC) at 0.025 µg/mL resulted in a significant reduction in micronuclei formation induced by MMC at concentrations of 75 and 150 µg/mL, suggesting antigenotoxic activity likely associated with the phytochemical constituents presented in the extract. Full article

Background/Objectives: Macrophage phenotype and function are highly sensitive to environmental cues; however, most in vitro studies rely on 2D culture systems that lack physiologically relevant structural context. The spatial dimensionality can influence immune cell signaling, yet the roles of these cells in regulating macrophage behavior remain incompletely understood. This study aimed to investigate how cultural dimensionality affects the phenotype, signaling, and functional activity of monocyte-derived macrophages. Methods: GFP-expressing THP-1 monocytes were differentiated into M0, M1, and M2 macrophages and cultured either on planar substrates or within 3D matrices composed of Matrigel or type I collagen. Macrophage morphology and viability were monitored. Membrane receptor expression and secreted cytokines were examined and quantified. Functional activity was further assessed through coculture experiments with RFP-expressing MDA-MB-231 breast cancer cells. Results: Compared with 2D culture, 3D environments induced distinct morphological and viability changes in macrophages. Collagen matrices supported sustained growth, subtype-specific morphologies, and enhanced functional activity, whereas Matrigel promoted aggregation and reduced viability. Core lineage markers remained stable across conditions, but activation-associated receptors and cytokine profiles were strongly influenced by dimensionality. 3D culture enhanced TNF-α expression and altered serglycin glycosylation patterns. In coculture assays, macrophage effects on tumor cell growth depended on polarization state and were more pronounced in 3D systems. Conclusions: These findings demonstrate that culture dimensionality and ECM composition are key regulators of macrophage phenotype and function. Collagen-based 3D systems better reproduce physiologically relevant macrophage behaviors than conventional 2D platforms, highlighting the value of structurally biomimetic models for immunological studies and therapeutic screening. Full article

Epimedium brevicornu is an important medicinal plant in China, whose main bioactive components are flavonoid glycosides. UDP-glycosyltransferases (UGTs) play key roles in flavonoid glycosylation and metabolic diversification. In this study, transcriptome data from four representative production regions in Gansu Province were used to systematically identify and analyze the UGT gene family in E. brevicornu. A total of 359 UGT members were identified, and 168 homologous genes with clear expression evidence were obtained from four geographical populations. Molecular evolutionary analysis showed that most UGT genes were under purifying selection, whereas UGT2, UGT52, UGT57, UGT241, UGT269, and UGT271 exhibited significant signals of positive selection in specific lineages (p < 0.05). Protein interaction analysis indicated that many UGT proteins were closely associated with key enzymes involved in flavonoid biosynthesis, including CHS (TT4), CHI (TT5), F3H, FLS, and DFR, suggesting their potential involvement in flavonoid metabolism. Promoter analysis further revealed a high enrichment of ERF (11,169 occurrences) and MYB (7673 occurrences) transcription factor binding sites in the upstream regions of UGT genes. In addition, UGT57 and UGT241 showed significantly higher expression levels in the QLH population. Molecular docking analysis indicated relatively strong binding affinities with quercetin, with binding energies of −7.23 kcal/mol and −4.62 kcal/mol, respectively. These results suggest that the sequence variation and differential expression of UGT genes may be associated with flavonoid glycosylation and ecological adaptation in Epimedium. This study provides a basis for understanding the evolutionary characteristics and expression patterns of the UGT gene family and offers candidate genes for future studies on flavonoid metabolism. Full article

Antimicrobial peptides (AMPs) have re-emerged as promising anti-infective agents, particularly against multidrug-resistant bacteria; however, their therapeutic development remains constrained by proteolytic degradation, host cell toxicity, and rapid systemic clearance. Rather than focusing solely on sequence discovery, recent efforts have shifted toward structural and supramolecular modification strategies aimed at improving stability, selectivity, and pharmacological performance. This review critically analyzes intramolecular modifications—including phosphorylation, glycosylation, acetylation, methylation, and backbone cyclization—that modulate peptide conformation and resistance to enzymatic degradation. In parallel, extramolecular approaches such as PEGylation, lipidation, and conjugation to antibiotics, siderophores, or antibodies are examined in the context of enhanced targeting and prolonged bioavailability. Particular emphasis is placed on localized delivery systems, including hydrogels, polymeric films, and nanofibrous scaffolds, which enable spatially controlled administration and mitigate systemic exposure. By integrating evidence from ex vivo and in vivo infection models, this work delineates the translational potential and remaining bottlenecks of chemically engineered AMP platforms for skin and soft tissue infections. Full article

Seasonal human coronaviruses (HCoVs) continue to undergo adaptive evolution under structural and immune-mediated constraints. We investigated the molecular epidemiology and spike (S) protein structural variation of circulating coronaviruses in Riyadh, Saudi Arabia, during the 2025–2026 winter season, with particular emphasis on genotype persistence and glycosylation architecture in HCoV-OC43. Among 293 nasopharyngeal aspirates (NPAs) collected from hospitalized patients with acute respiratory illness, HCoV-OC43 was detected in 26 cases (8.87%), whereas other seasonal coronaviruses were not identified. Partial sequencing of the S gene revealed 97.84–98.23% nucleotide identity relative to the prototype strain VR-759, with amino acid substitutions distributed at discrete positions rather than within extended variable domains, indicating structural conservation. Phylogenetic reconstruction demonstrated that all Riyadh isolates clustered within genotype C, together with previously circulating local strains, supporting sustained endemic persistence and in situ evolution. In silico analysis of the S protein glycosylation landscape identified four invariant N-linked glycosylation motifs (N-X-S/T) at residues 46, 121, 134, and 190, reflecting strong structural constraints on glycan-dependent folding and antigenic configuration. A genotype-associated K68N substitution generated an additional N-glycosylation motif (68NGTD) in multiple Riyadh isolates, potentially modifying local glycan shielding without disrupting the overall glycosylation framework. The preservation of core glycosylation sites alongside selective motif acquisition suggests evolutionary fine-tuning of S surface topology rather than large-scale structural remodeling. Collectively, these findings indicate that genotype C persistence in Riyadh is accompanied by conserved S architecture and subtle glycosylation adjustments that may modulate immune recognition while maintaining structural integrity. Continued high-resolution molecular surveillance will be critical for defining the functional consequences of S microevolution in endemic HCoVs. Full article

Astaxanthin (AST), despite its high bioactivity, exhibits poor stability and low bioavailability due to its strong lipophilicity and inherent degradation susceptibility. To overcome such a challenge, we developed a food-grade oleogel delivery system using a soy protein–arabinoxylan (SA) glycosylated complex modulated by different concentrations (0.5–3%) of sucrose ester (SE) or soy lecithin. We show that the emulsifier concentration has a non-linear effect on the oleogel microstructure: an optimal level of 1% had a significant impact on the interfacial compactness and network density, giving rise to improved thermal stability, rheological strength and AST encapsulation efficiency (81.27%). During in vitro digestion, the SA matrix in combination with emulsifiers allowed gastric protection and intestinal-targeted release of AST with a bioaccessibility of up to 88.84% (SAO-SE-AST). This controlled release profile directly translated into enhanced in vivo antioxidant efficacy in wild-type Bristol N2 Caenorhabditis elegans, as evidenced by reduced lipofuscin accumulation, elevated thermotolerance (survival rate: 64.44–73.33%), suppressed reactive oxygen species levels and activation of endogenous antioxidant enzymes (superoxide dismutase as well as glutathione peroxidase). Collectively, this research has uncovered that food-grade emulsifiers are not only stabilizers, but also key regulators of oleogel architecture and bioactive functionality. These results provide a structure–digestion–bioactivity correlation for protein–polysaccharide oleogels, representing a rational design strategy for high-performance delivery systems of lipid-soluble nutraceuticals. Full article

N-glycosylation is a fundamental post-translational modification that contributes to protein folding, stability, and secretion in eukaryotes. The catalytic subunit STT3 of the oligosaccharyltransferase complex mediates the transfer of preassembled oligosaccharides to nascent polypeptides in the endoplasmic reticulum. Here, we identified and functionally characterized PsSTT3B, one of the STT3 paralogs in Phytophthora sojae (P. sojae). PsSTT3B plays an important role in the growth, development, and pathogenicity of P. sojae. CRISPR/Cas9-mediated deletion of PsSTT3B resulted in reduced vegetative growth, sporangia production, and zoospore production in P. sojae. PsSTT3B deletion mutants demonstrated significantly reduced virulence on soybean leaves and etiolated seedlings. Importantly, PsSTT3B deletion mutants also exhibited reduced zoospore germination and diminished chemotaxis toward soybean isoflavones. Moreover, deletion of PsSTT3B increased sensitivity to tunicamycin and dithiothreitol and influenced the ConA-binding glycoprotein profile. Our findings show that PsSTT3B is associated with asexual development, virulence, and sensitivity to ER stress-related conditions of P. sojae. Our study suggests that PsSTT3B represents a potential candidate gene for the prevention and control of P. sojae. Full article

Fabrazyme (agalsidase beta) is a therapeutic enzyme whose clinical efficacy is contingent upon its complex N-glycosylation patterns. Nevertheless, comprehensive glycosylation profiling remains challenging due to high site-specific heterogeneity. To address this, three orthogonal liquid chromatography–mass spectrometry (LC-MS) approaches were employed: (1) released N-glycan analysis with fluorescence detection and MS annotation, (2) site-specific glycopeptide mapping, and (3) intact protein MS. The released glycan profiling method was validated for reproducibility, intermediate precision, and inter-laboratory transferability, thereby enabling reliable separation and quantification of neutral, phosphorylated, and sialylated species. Glycopeptide mapping revealed distinct site-specific distributions: N108 was found to predominantly carry sialylated complex glycans; N161 was enriched in phosphorylated oligomannose structures; and N184 displayed the highest heterogeneity, including bisphosphorylated and sialylated glycans. Intact protein analysis was performed on both intact and desialylated Fabrazyme, thereby enabling confirmation of glycan assignments. Desialylation reduced spectral complexity, thereby facilitating accurate mass matching with a combinatorial library generated from glycopeptide-level data. The complementary use of these three analytical levels provides a comprehensive view of Fabrazyme glycosylation, offering a robust reference for quality control and biosimilar development. Full article

Pinoresinol diglucoside (PDG), one of the major lignans isolated from E. ulmoides Oliver bark, has various pharmacological functions, including antihypertension and prevention of osteoporosis. However, the glycosyltransferase-encoding gene (GT) involved in regulating the glycosylation of pinoresinol to form PDG has not been reported in E. ulmoides. In this study, we screened and cloned the EuGT8 gene from E. ulmoides based on our transcriptome data. The expression pattern of the EuGT8 gene exhibited a strong positive correlation with dynamic changes in the PDG contents in three different organs of E. ulmoides. The expression level of the EuGT8 gene and PDG content were significantly decreased in asODN-EuGT8-treated shoot tips in comparison with the control group. Prokaryotic expression of the EuGT8 gene revealed that the purified EuGT8 protein could catalyze the conversion of pinoresinol into PDG. In addition, we performed transcriptional and metabolomic analyses to compare the differences between transgenic Arabidopsis and WT plants. A total of 1799 DEGs and 294 DEMs were identified in transgenic and WT plants. KEGG enrichment analysis showed that the DEGs were mainly enriched in phenylpropanoid biosynthesis, secondary metabolite biosynthesis, and starch/sucrose metabolism pathways. The DEMs were mainly enriched in ABC transporters, aminoacyl-tRNA biosynthesis, biosynthesis of amino acids, phenylpropanoid biosynthesis, and flavone and flavonol biosynthesis pathways. Correlation analysis between DEGs and DEMs identified a total of 231 DEGs associated with 38 DEMs, which were mainly distributed in multiple metabolic pathways. This finding provides both theoretical insights and genetic resources for breeding high-PDG E. ulmoides varieties, facilitating marker-assisted selection (MAS) and promoting sustainable E. ulmoides production in Guizhou. Full article