Search Results (210)

Human DNA dioxygenase ABH2 is a key enzyme of the AlkB family of Fe(II)/α-ketoglutarate-dependent oxygenases, which is specialized in removing alkyl groups from damaged DNA bases in the cell nucleus. At the same time, the occurrence of single-nucleotide polymorphisms (SNPs) in the human ABH2 gene can lead to amino acid substitutions that, in turn, may disrupt the normal functioning of the ABH2 enzyme. Currently, databases contain information about more than 2500 nucleotide substitutions in the ABH2 gene. Using a comprehensive bioinformatics approach, in this review, we analyzed over 200 non-synonymous ABH2 SNPs with eleven prediction programs to identify variants capable of negatively affecting its enzymatic activity. The combination of various programs with different evaluation algorithms and scoring approaches allows us to more reliably identify potentially deleterious amino acid substitutions. Moreover, the differences between the programs used allowed for comparison of their tendency to predict amino acid substitutions as deleterious. Structural analysis of the ABH2-substrate complex showed that selected functionally significant SNPs often affect the organization of the active site, reduce the efficiency of substrate binding, and/or disrupt the coordination of Fe²⁺ and α-ketoglutarate cofactors, leading to changes in catalytic efficiency. The data obtained from the conducted analysis suggest that naturally occurring polymorphisms in the ABH2 gene found in the human population may reduce the repair efficiency of DNA dioxygenase ABH2 and, consequently, modulate susceptibility to oncogenesis and influence the effectiveness of antitumor therapy for carriers of these SNPs. Full article

Recently, a growing interest has been focused to the role of vitamin C in chronic diseases. Type 2 Diabetes Mellitus and the Metabolic Syndrome are complex, chronic disorders intrinsically linked by a common underlying element, such as chronic low-grade inflammation and excessive oxidative stress. Vitamin C, or ascorbic acid, is an essential water-soluble micronutrient and a highly potent non-enzymatic antioxidant that is critical for scavenging reactive oxygen species and maintaining cellular redox balance. It represents a cofactor for many enzymes, being involved in many biological functions, such as normal immune system functioning, catecholamine metabolism, dietary iron absorption, and collagen biosynthesis. Individuals with type 2 diabetes mellitus and metabolic syndrome frequently exhibit lower circulating and dietary vitamin C levels compared to healthy controls, a deficiency that may be associated with disease-related inflammation and higher body weight. In this sense, it has been shown that vitamin C improves skeletal muscle insulin sensitivity in experimental settings and modulates critical functions like vascular endothelial health. However, this potential is challenged by the fact that chronic hyperglycemia can interfere with the active cellular uptake and transport of vitamin C, potentially leading to relative intracellular deficiency in diabetic patients regardless of intake. It is interesting to note that different studies have demonstrated an inverse relationship between vitamin C concentrations and the prevalence of metabolic syndrome and type 2 diabetes. Vitamin C supplementation in people with diabetes and metabolic syndrome has controversial effects. While several studies indicate a significant reduction in fasting blood glucose or HbA1c, others revealed no significant effect on insulin resistance. This review aims to explore the pathophysiological role and therapeutic potential of vitamin C in type 2 diabetes and metabolic syndrome. Full article

Dark fermentation of food waste for biohydrogen production can simultaneously achieve waste resource utilization and clean energy production. However, the widespread application of this technology remains constrained by challenges such as low substrate hydrolysis efficiency and suboptimal metabolic performance of functional microorganisms. This study evaluated the synergistic enhancement of biohydrogen production from food waste through dark fermentation by integrating thermal–alkaline (TA) pretreatment with varying concentrations (50, 100, 150, and 200 mg/L) of nickel–cobalt oxide nanoparticles (NiCo₂O₄ NPs), and the underlying mechanisms involved were systematically elucidated. The results demonstrated that individual TA pretreatment (pH 11, 70 °C, 1 h) and TA coupled with NiCo₂O₄ NPs (100 mg/L) significantly (p < 0.01) enhanced the cumulative biohydrogen yields of the food waste dark fermentation by 20.89% and 35.76%, respectively. Mechanism research revealed that TA pretreatment effectively facilitated the dissolution and hydrolysis of macro-molecular organics such as polysaccharides and proteins, thereby enhancing the bio-accessibility of fermentation substrates. The introduction of NiCo₂O₄ NPs further intensified the microbial biohydrogen-producing metabolism by augmenting enzymatic activity and enriching functional bacteria. NiCo₂O₄ NPs significantly (p < 0.001) enhanced the overall activity of hydrogenase by 95.10% compared to the control group (CG) by providing the cofactor of hydrogenase and accelerating electron transfer. Additionally, this synergistic strategy significantly (p < 0.01) increased the activities of hydrolases (e.g., protease and α-glucosidase), as well as key enzymes in acetate-type and butyrate-type fermentation pathways (e.g., acetate kinase and butyrate kinase), and enriched the biohydrogen-producing microbial community centered on Clostridium_sensu_stricto_1. This study systematically elucidated the synergistic strategy of TA pretreatment and NiCo₂O₄ NPs, which achieved dual-pathway reinforcement from substrate degradability to microbial metabolic activity. The findings are expected to provide theoretical support for developing efficient biohydrogen production technology from perishable organic solid waste. Full article

Magnesium (Mg²⁺) is the fourth most abundant cation in the human body and a critical cofactor in hundreds of enzymatic reactions that regulate energy metabolism, neuromuscular function, cardiovascular health, bone integrity, immune defense, and psychological well-being. Despite its essential roles, magnesium deficiency remains common worldwide, driven by inadequate dietary intake, chronic diseases, medication use, and lifestyle factors. Low magnesium status is associated with hypertension, type 2 diabetes, osteoporosis, migraines, depression, and chronic inflammation, whereas sufficient intake supports cardiometabolic resilience, skeletal strength, neurological stability, and healthy aging. This review synthesizes current evidence on magnesium metabolism, physiological functions, and the health consequences of deficiency, and it summarizes global status with attention to biomarker limitations, widespread suboptimal intake, and key demographic and lifestyle determinants. It also discusses dietary sources, supplementation, and innovative approaches such as food fortification, personalized nutrition, and improved diagnostic strategies. The evidence highlights magnesium as a modifiable factor with potential to lessen the burden of chronic diseases. Recognizing magnesium deficiency as a pressing but underappreciated public health issue, this article underscores the need for integrated strategies to optimize magnesium balance at both individual and population levels. Full article

The search for novel low-molecular regulators using molecular docking continues to be crucial for addressing challenges in modern biomedical science. However, the current literature lacks examples of modeling covalent interactions between the ligands being docked and those already present within the proteins, such as enzyme cofactors. This study aims to improve the existing algorithms for modeling such interactions, exemplified by those in thiamine diphosphate (ThDP)-dependent enzymes. Structures containing adducts of ThDP with enzyme substrates or inhibitors are used as protein templates; the putative ligand models are prepared as (R)- or (S)-hydroxy derivatives. The Gnina framework with AD4 or Vinardo favors ligand conformations resembling those found in the protein templates and consistent with their relative inhibitory potentials in experiments in vitro. For example, local hydrophobic regions within pyruvate and branched-chain 2-oxo acid dehydrogenase structures favor the binding of esterified substrate analogs compared to their de-esterified counterparts. The preferred binding of esterified vs. de-esterified ligands is absent or even reversed for 2-oxoglutarate dehydrogenase. As a result, covalent docking of 2-oxo acid analogs to enzyme structures containing ThDP coenzyme offers a predictive capability for protein–ligand complex formation and should be used when inhibitors mimic transition states in enzymatic reactions, as observed with ThDP-dependent catalysis. Full article

Phenol red is a widely used, low-cost, label-free colorimetric pH indicator that bridges traditional colorimetric assays with modern quantitative imaging and cell-based screening platforms. Its protonation-dependent absorbance shift (430–560 nm) allows for the real-time monitoring of extracellular acidification, which indirectly reflects cellular metabolism, growth, and respiration. Although phenol red lacks the molecular specificity of genetically encoded or fluorogenic biosensors, it remains useful in systems where pH changes are effective proxies for physiological processes. Existing tissue digestion protocols often overlook key parameters, especially pH control and enzyme cofactor use. This study presents a straightforward, spectrophotometric method to monitor and adjust the pH of low-volume (1 mL) buffered enzymatic dissociation media using phenol red and a plate reader. We titrated dissociation solutions to physiological pH (~7.4) using spectrophotometric pH measurements validated against conventional glass pH probe readings, confirming method reliability. Accurate pH assessment is critical for isolating viable primary cells for downstream applications such as tissue engineering, single-cell omics, and neurophysiological assays. We highlight that papain-based dissociation media supplemented with L-cysteine can be acidic (pH 6.6) if unadjusted, compromising cell viability. This accessible approach enhances reproducibility by promoting pH documentation concerning dissociation conditions that contribute to advancing consistency in biomedical, cellular, neuronal, and tissue engineering research. Full article

Biocatalysis is fundamental to biological processes and sustainable chemical productions. Over time, the biocatalysis strategy has been widely researched. Initially, biomanufacturing and catalysis of high-value chemicals were carried out through direct immobilization and application of biocatalysts, including natural enzymes and living cells. With the evolution of green chemistry and environmental concern, hybrid photoelectro-biocatalysis (HPEB) platforms are seen as a new approach to enhance biocatalysis. This strategy greatly expands the domain of natural biocatalysis, especially for bio-based components. The selective valorization of renewable furan derivatives, such as 5-hydroxymethylfurfural (HMF) and furfural, is central to advancing biomass-based chemical production. Biocatalysis offers high chemo-, regio-, and stereo-selectivity under mild conditions compared with traditional chemical catalysis, yet it is often constrained by the costly and inefficient regeneration of redox cofactors like NAD(P)H. Photoelectrocatalysis provides a sustainable means to supply reducing equivalents using solar or electrical energy. In recent years, hybrid systems that integrate biocatalysis with photoelectrocatalysis have emerged as a promising strategy to overcome this limitation. This review focuses on recent advances in such systems, where photoelectrochemical platforms enable in situ cofactor regeneration to drive enzymatic transformations of furan-based substrates. We critically analyze representative coupling strategies, materials and device configurations, and reaction engineering approaches. Finally, we outline future directions for developing efficient, robust, and industrially viable hybrid catalytic platforms for green biomass valorization. Full article

Background/Objectives: Vitamin C plays a vital role in human health, functioning as a powerful antioxidant and enzymatic cofactor. Although vitamin C bioavailability from food versus supplements has been debated, few studies have examined how intake form affects absorption and physiological markers. Methods: This randomized, controlled, crossover trial aimed to compare the bioavailability of vitamin C consumed as a supplement, through raw fruits and vegetables, or through fruit and vegetable juice. Twelve healthy adults underwent three 1-day crossover trials, each separated by a 2-week washout. Participants consumed 101.7 mg of vitamin C via powder, raw fruits and vegetables (186.8 g), or juice (200 mL). Plasma and urinary vitamin C concentrations, urinary metabolites (¹H NMR), and antioxidant activity (ORAC and TRAP) were assessed over 24 h. Results: All interventions elevated plasma vitamin C levels, with juice yielding the highest AUC (25.3 ± 3.2 mg/dL·h). Urinary vitamin C increased in all groups. Metabolomics revealed increased urinary excretion of mannitol, glycine, taurine, dimethylglycine (DMG), and asparagine, and decreased choline and dimethylamine (DMA). Notably, urinary mannitol increased only in the morning. Choline significantly decreased after powder intake (p = 0.001), with similar trends observed in the other groups. DMG and glycine increased following raw and juiced vegetable intake. Antioxidant activity showed transient ORAC elevation post-powder but no sustained improvements. Conclusions: Vitamin C is bioavailable from all intake forms, with juice providing the most efficient absorption. Urinary metabolite changes suggest microbiota-related modulation, while antioxidant activity improvements were limited. Full article

Specific expression of genes is fundamental for defining the identity and the functional state of cells. Sequence-specific transcription factors interpret the information contained in DNA sequence motifs and recruit cofactors to modify chromatin and control RNA polymerases. This multi-step process typically involves several transcription factors and cofactors with different enzymatic activities. Post-translational modifications (PTMs) of histones are one key mechanism to control chromatin structure and polymerase activity and thus gene transcription. The methylation of histone H3 at lysine 4 (H3K4) is a modification of accessible chromatin, including enhancers and promoters, and also sites of recombination and some forms of DNA damage. H3K4 methylation is catalyzed by six lysine methyltransferase complexes, referred to as KMT2 or COMPASS-like complexes. These are important in processes related to transcription and contribute to recombination in T and B cells. PRDM9 and ASH1L are H3K4 methyltransferases involved in meiotic recombination and DNA repair, respectively. In transcription, H3K4 mono- and tri-methylation are located at enhancers and promoters, respectively. These modifications, either alone or in combination with other histone PTMs, provide binding sites for transcriptional cofactors. Through these sites, H3K4 methylation affects chromatin accessibility and histone PTMs, typically resulting in a favorable environment for transcription. H3K4 tri-methylation also recruits and regulates RNA polymerase II (RNAPII) complexes, which interact with KMT2 complexes, generating positive feedforward loops to promote transcription. Thus, H3K4 methylation has broad activities that are key to different chromatin-associated processes. Full article

Zinc (Zn) is an essential micronutrient for plant growth, serving as a co-factor in enzymatic processes and pigment biosynthesis. In horticultural crops such as lettuce, Zn fertilization is increasingly relevant for optimizing yield and nutritional quality. In this study, a greenhouse pot experiment was conducted using Lactuca sativa L. cv. Romana Verano (Ramiro Arnedo) to evaluate the effects of four Zn sources with contrasting physio-chemical properties—ZnSO₄, a synthetic chelate containing DTPA, EDTA, and HEDTA, a Zn–lignosulphonate complex, and ZnO nanoparticles—applied to soil at rates of 15, 30, 60, and 120 mg Zn·kg⁻¹. Morphometric traits, photosynthetic pigmentation, and photosystem performance were assessed to determine differences in plant response. Results showed that low to moderate Zn supply (15–60 mg Zn·kg⁻¹) maintained growth, leaf number, stem diameter, and biomass without significant changes compared to the control. In contrast, the highest dose (120 mg Zn·kg⁻¹), particularly in chelated forms, led to reductions in growth and yield exceeding 80%, reflecting supra-optimal effects. Although lignosulphonate and nanoparticles sources lowered soil Zn availability, they did not affect lettuce growth or yield, indicating their potential as safer agricultural alternatives to conventional Zn fertilizers. Photosynthetic efficiency, measured through chlorophyll fluorescence and electron transport activity, was positively modulated by adequate Zn levels but declined at excessive concentrations. These findings highlight that Zn efficiency strongly depends on its chemical form and applied dose, providing practical insights for optimizing Zn fertilization strategies in lettuce and other horticultural crops. Full article

This study reports on the direct electron transfer (DET) ability of the enzyme spermidine dehydrogenase (SpDH) and its use in a DET-type enzymatic sensor for detecting spermine. SpDH was found to exhibit internal electron transfer from its cofactor, flavin adenine dinucleotide (FAD), to heme b. This was confirmed by observing the heme b-derived reduction peak at 560 nm in the presence of spermine, the substrate. SpDH was immobilized on a gold electrode via a dithiobis (succinimidyl hexanoate) self-assembled monolayer. The cyclic voltammetry analysis of the SpDH-immobilized gold electrode revealed an increased oxidation current in the presence of 0.1 mM spermine with an onset potential of −0.14 V vs. Ag/AgCl in the absence of an additional external electron acceptor. This result confirmed that SpDH is capable of DET. Chronoamperometric analyses were conducted using an SpDH-immobilized gold electrode with spermine as the substrate under a 0 V oxidation potential vs. Ag/AgCl using an artificial saliva matrix containing 10 µM ascorbic acid and 100 µM uric acid. The sensor exhibited good linear correlation between the current increase and spermine concentration from 0.2 to 2.0 µM, with a limit of detection of 0.084 µM, which encompasses the physiologically relevant spermine concentration found in the saliva. Primary structure alignments and 3D structure predictions revealed that all SpDH homologs possess two conserved histidine residues in the same location on the surface as the heme b ligand of SpDH. This indicates their potential for DET-ability with an electrode. Full article

Bacterial inclusion bodies (IBs) are still generally considered to be waste products of recombinant protein production, despite various studies that have challenged this conventional view in the last two decades, and have been proposed for use as immobilized enzymes in vivo for biocatalysis. Current advances in genetic and molecular biology make it possible to perform multienzymatic reactions or enzymatic cascades to synthesize valuable products. When cascades need cofactor regener tion, it is difficult to use “cheap” whole cells or their lysates, and “expensive” enzyme purification is required. The capture of enzymatic activity into active IBs (aIBs), well-separable protein aggregates from cell lysate, could represent a usable compromise between purified enzymes and cell lysates. It is shown here that the combination of two polyphosphate kinases (PPKs) in the form of aIBs leads to almost 10-fold ATP regeneration and 100% UTP utilization without degradation into adenosine or uridine. PPKs have been combined with N-acetylhexosamine 1-kinase and N-acetylglucosamine-1-phosphate uridyltransferase to produce valuable UDP-N-acetylglucosamine, but the described approach could be used in various multienzymatic syntheses to avoid enzyme purification and ensure nucleotide triphosphate regeneration. Full article

Photocatalytic nitrogen fixation under ambient conditions offers a sustainable alternative to the energy-intensive Haber–Bosch process, yet remains limited by the inertness of N≡N bonds and sluggish multi-electron/proton transfer kinetics. Nature’s nitrogenase enzymes, featuring the FeMo cofactor and ATP-driven electron cascades, inspire a new generation of artificial systems capable of mimicking their catalytic precision and selectivity. This review systematically summarizes recent advances in bio-inspired photocatalytic nitrogen reduction, focusing on six key strategies derived from enzymatic mechanisms: Fe–Mo–S active site reconstruction, hierarchical electron relay pathways, ATP-mimicking energy modules, defect-induced microenvironments, interfacial charge modulation, and spatial confinement engineering. While notable progress has been made in enhancing activity and selectivity, challenges remain in dynamic regulation, mechanistic elucidation, and system-level integration. Future efforts should prioritize operando characterization, adaptive interface design, and device-compatible catalyst platforms. By abstracting nature’s catalytic logic into synthetic architectures, biomimetic photocatalysis holds great promise for scalable, green ammonia production aligned with global decarbonization goals. Full article

Background and Methods: To address the need for dietary interventions in sub-healthy populations and promote sustainable utilization of agricultural by-products, we isolated Pinus pumila hypoglycemic peptide (PHP) from nut oil meal through enzymatic extraction, ion exchange and gel chromatography purification, and simulated gastric digestion. Results: PHP exhibited significant inhibitory activity against α-amylase and α-glucosidase. In type 2 diabetic mice, PHP significantly ameliorated the “three-more-one-less” syndrome, reduced glycosylated hemoglobin and insulin levels, mitigated liver and kidney tissue lesions, and improved glucose and lipid metabolic disorders—effects partly supported by its enhancement of intestinal barrier function via restoring gut microbiota diversity. Gut microbiota analysis revealed that PHP exerts hypoglycemic effects by regulating gut microbial composition: increasing SCFA-producing taxa, reducing pro-inflammatory/metabolic disorder-associated taxa, and normalizing the Firmicutes/Bacteroidetes ratio. KEGG pathway analysis demonstrated that PHP mediates synergistic hypoglycemic effects by regulating carbohydrate metabolism, amino acid metabolism, and cofactor/vitamin metabolism. Conclusions: This work provides a theoretical foundation for developing natural functional foods from agricultural by-products, supporting PHP’s potential as a dietary supplement for metabolic regulation. Full article

Understanding the structure–function relationships of pyridoxal-5′-phosphate (PLP)-dependent transaminases is key to advancing pyridoxal-phosphate-dependent catalysis and engineering transaminases for industrial applications. Despite our extensive knowledge of PLP-dependent enzymatic reactions, engineering transaminase activity and stability remains challenging. Here, we present the functional characterization of a novel PLP-dependent fold type IV transaminase from Desulfomonile tiedjei, alongside a detailed analysis of PLP binding and holoenzyme stability. This new transaminase exhibits activity toward various D-amino acids and (R)-phenylethylamine. Structural modeling and site-directed mutagenesis of residues in the second shell of the PLP-binding site revealed their roles in cofactor binding and the transaminase’s catalytic efficiency. Notably, the T199Q variant demonstrated a fivefold increase in PLP affinity and improved activity under alkaline conditions. This is attributed to a newly formed hydrogen bond that stabilizes the N1-binding region of PLP. Glutamine at position 199 is not observed in homologous transaminases, making this non-natural substitution a novel and beneficial modification. These findings emphasize the importance of second-shell interactions in stabilizing PLP and expand our understanding of the structural diversity within PLP fold type IV transaminases. This paves the way for the engineering of more stable and versatile biocatalysts for industrial applications. Full article