Search Results (169)

Background: Neonatal seizures are critical neurological events with long-term implications for brain development. Standard antiseizure medications, such as phenobarbital, often yield suboptimal seizure control and may be associated with neurotoxicity. This narrative review explores the role of vitamin B6 as a precision therapy in neonatal seizure syndromes, particularly in pyridoxine-responsive conditions. Methods: We conducted a narrative review of the biochemical functions of vitamin B6, focusing on its active form, pyridoxal 5′-phosphate (PLP), and its role as a coenzyme in neurotransmitter synthesis. We examined the genetic and metabolic disorders linked to vitamin B6 deficiency, such as mutations in pyridox(am)ine 5’-phosphate oxidase (PNPO), Aldehyde Dehydrogenase 7 Family Member A1 (ALDH7A1), alkaline locus phosphatase (ALPL), and cystathionine β-synthase (CBS), and discussed the clinical rationale for empirical administration in acute neonatal seizure settings. Results: Vitamin B6 is essential for the synthesis of gamma-aminobutyric acid (GABA), dopamine, and serotonin, with PLP-dependent enzymes such as glutamic acid decarboxylase and aromatic L-amino acid decarboxylase playing central roles. Deficiencies in PLP due to genetic mutations or metabolic disruptions can result in treatment-resistant neonatal seizures. Early supplementation, especially in suspected vitamin B6-dependent epilepsies, may provide both diagnostic clarity and seizure control, potentially reducing exposure to conventional antiseizure medications. Conclusions: Vitamin B6-responsive epilepsies highlight the clinical value of mechanism-based, individualized treatment approaches in neonatology. Incorporating genetic and metabolic screening into seizure management may improve outcomes and aligns with the principles of precision medicine. Full article

Cystathionine γ-lyase (CSE) is a pyridoxal 5′-phosphate (PLP)-dependent enzyme that catalyzes the final step of the transsulfuration pathway, converting cystathionine into cysteine. Additionally, CSE is also essential for the formation of cysteine hydropolysulfide (Cys-S-(S)n-H), known as supersulfides, by metabolizing cystine under pathological conditions. We previously reported that, during cystine metabolism, CSE undergoes self-inactivation through polysulfidation at the Cys136 residue. Here, contrary to the anticipated role of L-S-nitrosocysteine (L-CysNO) as a nitric oxide (NO) donor, we demonstrate that it serves as a substrate for CSE and that its metabolites inhibit the activity of the enzyme during L-CysNO metabolism. The in vitro incubation of CSE—but not the Cys136/171Val mutant—with L-CysNO resulted in the dose-dependent inhibition of supersulfide production, which was not reversed by the reducing agents. Notably, CSE activity remained unchanged upon preincubation with other NO donors, such as S-nitrosoglutathione or D-CysNO, but was inhibited when coincubated with cysteine. Furthermore, when PLP was removed from the CSE/L-CysNO premix, L-CysNO no longer inhibited CSE activity, suggesting that CSE metabolizes L-CysNO and that its metabolites contribute to enzyme inactivation. Indeed, we identified thionitrous acid and pyruvate as the primary CSE/L-CysNO reaction products. Thus, we establish L-CysNO as a CSE substrate and demonstrate that its metabolites act as enzyme inhibitors through a novel irreversible modification at the Cys136/171 residues. Full article

The ability of enzymes to access various conformational states is often essential for their catalytic activity. Lysine 5,6-aminomutase (5,6-LAM), a pyridoxal 5′-phosphate (PLP) and 5′-deoxyadenosylcobalamin (dAdoCbl)dependent enzyme, catalyzes 1,2-amino shift in lysine isomers by shuttling between an open conformational state and a closed conformational state. Nevertheless, suicide inactivation of 5,6-LAM is an obstacle to the realization of its potential as a biocatalyst. In this work, the fate of the reaction of 5-hydroxylysine, an analogue of lysine, is investigated using spectroscopic and computational methods. Although 5-hydroxylysine does not afford any product, results obtained from UV–visible and electron paramagnetic resonance (EPR) spectroscopies demonstrate that initial steps of the catalytic cycle are performed with it. Simulation of the weakly spin-coupled spectrum estimates an intermediate distance between the PLP substrate-based radical and Co(II) in comparison to the that in the open state and the closed state. This distinct conformational state, different from the open state and the closed state, is alluded to in its putative role in suicide inactivation and denoted as the suicidally-inactivated state. Our findings highlight the emergence of EPR spectroscopy as a powerful tool to uncover the hidden conformations in radical enzymes. These results provide new insights into the suicide inactivation of dAdoCbl-dependent enzymes. 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

Global food security relies on wheat, maize, and soybean, yet their cultivation faces escalating threats from Fusarium head blight (FHB) pathogens. We demonstrate that agricultural intensification enables cross-kingdom root infections by Fusarium graminearum and F. asiaticum across these crops. Screening of 180 Fusarium strains revealed tripartite host infectivity, with transcriptomics uncovering host-adapted virulence strategies. Transcriptome analysis identified distinct gene expression patterns during the infection of each crop, with F. graminearum employing host-specific genes, such as FgPPDT1 (a pyridoxal phosphate-dependent transferase), for maize root infection. The FgPPDT1 knockout mutant (Δfgppdt1) exhibited severely impaired root colonization. Our findings establish differential gene expression as a regulatory axis for cross-host adaptation, directly linking FHB transmission risks to wheat–maize intercropping and wheat-soybean rotations. Full article

This study aimed to assess the extent of vitamin B₁ and B₆ vitamer loss during a single peritoneal dialysis (PD) session using a combination of chromatographic techniques and chemometric analysis. Dialysis effluent samples were collected from 41 PD patients (22 on continuous ambulatory peritoneal dialysis (CAPD) and 19 on automated peritoneal dialysis (APD)) during a standardised peritoneal equilibration test. Concentrations of thiamine monophosphate, thiamine diphosphate (ThDP), pyridoxine, pyridoxal (PL), and pyridoxamine were determined using high-performance liquid chromatography with a fluorescence detector. The analytical method was validated in terms of sensitivity, linearity, accuracy, and recovery. Multiple regression analysis was employed to identify potential clinical and demographic predictors of vitamin washout. All vitamers except pyridoxal 5-phosphate (PLP) were detectable in dialysis effluents. ThDP exhibited the greatest loss among the B₁ forms (ca. 0.05–0.57 mg/24 h), while PL exhibited the most significant loss among the B₆ forms (ca. 0.01–0.19 mg/24 h). Vitamin losses varied depending on the dialysis modality (continuous ambulatory peritoneal dialysis, or CAPD, versus automated peritoneal dialysis, or APD) and the peritoneal transport category. Regression analysis identified body weight, haemoglobin, and haematocrit as independent predictors of ThDP washout (R² = 0.58). No statistically robust models were established for the other vitamers. Even short medical procedures (such as single PD) can result in measurable losses of water-soluble vitamins, particularly ThDP and PL. The results emphasise the importance of personalised vitamin supplementation for PD patients and suggest that body composition and haematological parameters significantly influence the loss of thiamine. Full article

Sarcopenia, the progressive loss of muscle mass, strength, and regenerative capacity with age, is driven by interconnected processes such as oxidative stress, chronic inflammation, mitochondrial dysfunction, and reduced activity of muscle stem cells. As the population ages, nutritional strategies that target these mechanisms are becoming increasingly important. This review focuses on nicotinamide (vitamin B3) and pyridoxine (vitamin B6), two essential micronutrients found in functional foods, which play complementary roles in redox regulation, immune balance, and muscle repair. Nicotinamide supports nicotinamide adenine dinucleotide (NAD⁺) metabolism, boosts mitochondrial function, and activates sirtuin pathways involved in autophagy and stem cell maintenance. Pyridoxine, via its active form pyridoxal 5′-phosphate (PLP), is key to amino acid metabolism, antioxidant defense, and the regulation of inflammatory cytokines. We summarize how these vitamins influence major molecular pathways such as Sirtuin1 (SIRT1), protein kinase B (AKT)/mechanistic target of rapamycin (mTOR), Nuclear factor-κB (NF-κB), and Nrf2, contributing to improved myogenic differentiation and protection of the aging muscle environment. We also highlight emerging preclinical and clinical data, including studies suggesting possible synergy between B3 and B6. Finally, we discuss how biomarkers such as PLP, nicotinamide mononucleotide (NMN), and C-reactive protein (CRP) may support the development of personalized nutrition strategies using these vitamins. Safe, accessible, and mechanistically grounded, nicotinamide and pyridoxine offer promising tools for sarcopenia prevention and healthy aging. Full article

Background/Objectives: We examined the association between dietary intake and blood concentrations of one-carbon metabolism (OCM)-related nutrients in the European Prospective Investigation into Cancer and Nutrition (EPIC). Methods: Blood concentrations and dietary intake of the vitamins riboflavin (B2), Pyridoxal 5′-phosphate (PLP and B6), folate (B9), B12, and methionine, concentrations of homocysteine, and dietary intake of betaine, choline, and cysteine were pooled from 16,267 participants in nine EPIC nested case–control studies. Correlation analyses between dietary intakes and blood concentrations were carried out. Principal component (PC) analysis identified latent factors in the two sets of measurements. Results: Pearson correlations between dietary intakes and blood concentrations ranged from 0.08 for methionine to 0.12 for vitamin B2, 0.15 for vitamin B12, 0.17 for vitamin B6, and 0.19 for folate. Individual dietary intakes showed higher correlations (ranging from −0.14 to 0.82) compared to individual blood concentrations (from −0.31 to 0.29). Correlations did not vary by smoking status, case–control status, or vitamin supplement use. The first PC of dietary intakes was mostly associated with methionine, vitamin B12, cysteine, and choline, while the first PC of blood concentrations was associated with folate and vitamin B6. Conclusions: Within this large European study, we found weak to moderate associations between dietary intakes and concentrations of OCM-related nutrients. Full article

Hypophosphatasia (HPP) is a rare inborn error of metabolism caused by pathogenic variants in ALPL, coding for tissue non-specific alkaline phosphatase. HPP patients suffer from impaired bone mineralization, and in severe cases from vitamin B₆-responsive seizures. To study HPP, we generated alpl^-/- zebrafish using CRISPR/Cas9 gene-editing technology. At 5 days post fertilization (dpf), no alpl mRNA and 89% lower total alkaline phosphatase activity was detected in alpl^-/- compared to alpl^+/+ embryos. The survival of alpl^-/- zebrafish was strongly decreased. Alizarin red staining showed decreased bone mineralization in alpl^-/- embryos. B₆ vitamer analysis revealed depletion of pyridoxal and its degradation product 4-pyridoxic acid in alpl^-/- embryos. Accumulation of d3-pyridoxal 5′-phosphate (d3-PLP) and reduced formation of d3-pyridoxal in alpl^-/- embryos incubated with d3-PLP confirmed Alpl involvement in vitamin B₆ metabolism. Locomotion analysis showed pyridoxine treatment-responsive spontaneous seizures in alpl^-/- embryos. Metabolic profiling of alpl^-/- larvae using direct-infusion high-resolution mass spectrometry showed abnormalities in polyamine and neurotransmitter metabolism, suggesting dysfunction of vitamin B₆-dependent enzymes. Accumulation of N-methylethanolaminium phosphate indicated abnormalities in phosphoethanolamine metabolism. Taken together, we generated the first zebrafish model of HPP that shows multiple features of human disease and which is suitable for the study of the pathophysiology of HPP and for the testing of novel treatments. Full article

As crucial vectors that transmit pathogens to humans and livestock, ticks pose substantial global health threats and economic burdens. We analyzed 328 tick genomes to explore the population’s genetic structure and the adaptive evolution of H. longicornis and R. microplus, two tick species with distinct life cycle characteristics. We observed distinct genetic structures in H. longicornis and R. microplus. Gene flow estimation revealed a closer genetic connection in R. microplus than H. longicornis, which was facilitated by geographical proximity. Notably, we identified a set of candidate genes associated with possible adaptations. Specifically, the immune-related gene DUOX and the iron transport gene ACO1 showed significant signals of natural selection in R. microplus. Similarly, H. longicornis exhibited selection in pyridoxal-phosphate-dependent enzyme genes associated with heme synthesis. Moreover, we observed significant correlations between the abundance of pathogens, such as Rickettsia and Francisella, and specific tick genotypes, which highlights the role of R. microplus in maintaining these pathogens and its adaptations that influence immune responses and iron metabolism, suggesting potential coevolution between vectors and pathogens. Our study highlights the vital genes involved in tick blood feeding and immunity, and it provides insights into the coevolution of ticks and tick-borne pathogens. Full article

Human cysteine desulfurase (NFS1) participates in numerous critical cellular processes, including iron–sulfur (Fe-S) cluster biosynthesis and tRNA thiolation. NFS1 overexpression has been observed in a variety of cancers, and thus it has been considered a promising anti-tumor therapeutic target. To date, however, no inhibitors targeting NFS1 have been identified. Here, we report the identification of the first potent small-molecule inhibitor (Compound 53, PubChem CID 136847320) of NFS1 through a combination of virtual screening and biological validation. Compound 53 exhibited good selectivity against two other pyridoxal phosphate (PLP)-dependent enzymes. Treatment with Compound 53 inhibited the proliferation of lung cancer (A549) cells (IC₅₀ = 16.3 ± 1.92 μM) and caused an increase in cellular iron levels due to the disruption of Fe-S cluster biogenesis. Furthermore, Compound 53, in combination with 2-AAPA, an inhibitor of glutathione reductase (GR) that elevates cellular reactive oxygen species (ROS) levels, further suppressed the proliferation of A549 cells by triggering ferroptotic cell death. Additionally, the key residues involved in the binding of the inhibitor to the active center of NFS1 were identified through a combination of molecular docking and site-directed mutagenesis. Taken together, we describe the identification of the first selective small-molecule inhibitor of human NFS1. Full article

ACS (1-aminocyclopropane-1-carboxylic acid synthase) is a member of the aminotransferase superfamily and a pyridoxal phosphate-dependent enzyme. ACS is also a rate-limiting enzyme for the biosynthesis of ethylene and has been linked with plant development, growth, and stress responses. However, information on ACS genes in the soybean genome is limited. In this study, we identified ACS genes in soybean through phylogenetic trees and conserved motifs and analyzed their cis-acting elements, subcellular localization, and expression patterns. Twenty-two members of the ACS family were identified in soybean, and they were divided into four subfamilies based on phylogenetic relationships. Moreover, the results of Arabidopsis mesophyll protoplasts showed that GmACS1, GmACS8, and GmACS15 were all localized in the nucleus and cell membrane. Cis-regulatory elements and qRT-PCR analyses indicated markedly increased levels of GmACS transcripts under hormone treatments and abiotic stress conditions (drought, alkalinity, and salt). In addition, under different abiotic stresses, the potential functional variations across the GmACS isoforms were mirrored in their differential expression. The analysis of transcriptional response to salinity indicated that salt stress might primarily be mediated by the GmACS15 gene. GmACS15 was also found to reduce salt-induced oxidative damage by modulating the ROS-scavenging system, cellular redox homeostasis, and maintaining intracellular Na⁺/K⁺ balance. The results of this investigation revealed the involvement of the ACS gene family in soybean stress-response pathways, including the identification of a potential target for enhancing salt tolerance in soybean. Full article

Background/Objectives: Antibiotic-resistant Staphylococcus aureus represents a growing threat in the modern world, and new antibiotic targets are needed for its successful treatment. One such potential target is the pyridoxal-5′-phosphate (PLP)-dependent cysteine desulfurase (SaSufS) of the SUF-like iron–sulfur (Fe-S) cluster biogenesis pathway upon which S. aureus relies exclusively for Fe-S synthesis. The current methods for measuring the activity of this protein have allowed for its recent characterization, but they are hampered by their use of chemical reagents which require long incubation times and may cause undesired side reactions. This problem highlights a need for the development of a rapid quantitative assay for the characterization of SaSufS in the presence of potential inhibitors. Methods: A spectrophotometric assay based on the well-documented absorbance of PLP intermediates at 340 nm was both compared to an established alanine detection assay and used to effectively measure the activity of SaSufS incubated in the absence and presence of the PLP-binding inhibitors, D-cycloserine (DCS) and L-cycloserine (LCS) as proof of concept. Methicillin-resistant S. aureus strain LAC was also grown in the presence of these inhibitors. Results: The Michaelis–Menten parameters k_cat and K_m of SaSufS were determined using the alanine detection assay and compared to corresponding intermediate-based values obtained spectrophotometrically in the absence and presence of the reducing agent tris(2-carboxyethyl)phosphine (TCEP). These data revealed the formation of both an intermediate that achieves steady-state during continued enzyme turnover and an intermediate that likely accumulates upon the stoppage of the catalytic cycle during the second turnover. The spectrophotometric method was then utilized to determine the half maximal inhibitory concentration (IC₅₀) values for DCS and LCS binding to SaSufS, which are 2170 $\pm$ 920 and 62 $\pm$ 23 μM, respectively. Both inhibitors of SaSufS were also found to inhibit the growth of S. aureus. Conclusions: Together, this work offers a spectrophotometric method for the analysis of new inhibitors of SufS and lays the groundwork for the future development of novel antibiotics targeting cysteine desulfurases. Full article

Background/Objectives: Septic cardiomyopathy (SCM) is a severe cardiac complication of sepsis, characterized by cardiac dysfunction with limited effective treatments. This study aimed to identify repurposable drugs for SCM by integrated multi-omics and network analyses. Methods: We generated a mouse model of SCM induced by lipopolysaccharide (LPS) and then obtained comprehensive metabolic and genetic data from SCM mouse hearts using ultra-performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) and RNA sequencing (RNA-seq). Using network proximity analysis, we screened for FDA-approved drugs that interact with SCM-associated pathways. Additionally, we tested the cardioprotective effects of two drug candidates in the SCM mouse model and explored their mechanism-of-action in H9c2 cells. Results: Network analysis identified 129 drugs associated with SCM, which were refined to 14 drug candidates based on strong network predictions, proven anti-infective effects, suitability for ICU use, and minimal side effects. Among them, acetaminophen and pyridoxal phosphate significantly improved cardiac function in SCM moues, as demonstrated by the increased ejection fraction (EF) and fractional shortening (FS), and the reduced levels of cardiac injury biomarkers: B-type natriuretic peptide (BNP) and cardiac troponin I (cTn-I). In vitro assays revealed that acetaminophen inhibited prostaglandin synthesis, reducing inflammation, while pyridoxal phosphate restored amino acid balance, supporting cellular function. These findings suggest that both drugs possess protective effects against SCM. Conclusions: This study provides a robust platform for drug repurposing in SCM, identifying acetaminophen and pyridoxal phosphate as promising candidates for clinical translation, with the potential to improve treatment outcomes in septic patients with cardiac complications. Full article

The first monomeric pyridoxal-5′-phosphate (PLP)-dependent transaminase from a marine, aromatic-compound-degrading, sulfate-reducing bacterium Desulfobacula toluolica Tol2, has been studied using structural, kinetic, and spectral methods. The monomeric organization of the transaminase was confirmed by both gel filtration and crystallography. The PLP-dependent transaminase is of the fold type IV and deaminates D-alanine and (R)-phenylethylamine in half-reactions. The enzyme shows high stereoselectivity; no deamination of L-amino acids and (S)-phenylethylamine is detected. Structural analysis and subsequent mutagenesis led to the conclusion that the monomeric architecture of the enzyme is the only one possible and sufficient for stereoselectivity and PLP binding, but not for the overall double-substrate transamination reaction and the stability of the holo form with the reduced cofactor—pyridoxamine-5′-phosphate. These results extend the structural university of the PLP fold type IV enzymes and demonstrate the need for deeper analysis of the sequence–structure–function relationships in the transaminases. Full article