Glucose-6-phosphate Dehydrogenase Research Articles

Fra-1 affects chemotherapy sensitivity by inhibiting ferroptosis in gastric cancer cells

Aim: Gastric cancer (GC) is one of the common malignant tumors, and most patients with advanced GC often develop chemotherapy resistance, resulting in poor chemotherapy efficacy. Therefore, it is crucial to clarify the specific mechanisms of their chemotherapy resistance. Methods: In this study, we analyzed the correlation between fos-related antigen-1 (Fra-1) and chemotherapy resistance in GC using bioinformatics, cell counting kit-8 (CCK8), and 5-ethynyl-2’-deoxyuridine (EDU) combined with flow cytometry; furthermore, we used energy metabolomics sequencing, combined with ChIP-qPCR technology, to elucidate the specific role of Fra-1 in chemotherapy resistance of GC cells and its related mechanisms. Results: We found that high Fra-1 expression was closely related to chemotherapeutic drugs in GC cells, as demonstrated by bioinformatics analysis combined with EDU and CCK8 experiments. Energy metabolomics combined with in vitro cellular experimental analysis revealed that the pentose phosphate pathway (PPP) was activated in GC cells with high Fra-1 expression, along with an increase in the synthesis of metabolites such as nicotinamide adenine dinucleotide phosphate (NADPH) and glutathione (GSH), a decrease in the level of reactive oxygen species (ROS), and the inhibition of their ferroptosis. In addition, ChIP-qPCR experiments confirmed that Fra-1 binds to the promoter of glucose-6-phosphate dehydrogenase (G6PD), a key rate-limiting enzyme of the PPP, and transcriptionally regulates its expression, which in turn activates the PPP and promotes chemotherapy resistance in GC cells. Conclusion: Our research findings suggest that Fra-1 activates the PPP by upregulating G6PD transcriptional activity and inhibiting its ubiquitination level, inhibiting ferroptosis in GC cells and inducing chemoresistance. This provides an experimental basis for screening potential molecular targets for chemotherapy resistance in GC patients.

Activities of Energy and Carbohydrate Metabolism Enzymes in Rainbow Trout Оncorhynchus mykiss Walb. upon Introduction of 24-hour Lighting in Aquaculture in Southern Russia.

Activities of energy and carbohydrate metabolism enzymes (cytochrome c oxidase (COX), pyruvate kinase (PK), glucose 6-phosphate dehydrogenase (G6PD), glycerol 1-phosphate dehydrogenase (G1PDH), lactate dehydrogenase (LDH), and aldolase) were studied in rainbow trout Oncorhynchus mykiss Walb. fish grown in aquaculture in North Ossetia-Alania after introducing a regime with 24-h lighting and night feeding. COX and PK activities in the liver of fish from the experimental group were found to be significantly higher than in control fish, indicating an increase in aerobic ATP synthesis. Aldolase activity in organs of fish grown with 24-h lighting was lower than in the control fish, indicating a decrease in carbohydrate utilization in glycolysis in muscles and a lower intensity of gluconeogenesis in the liver. The differences made it possible to assume that the introduction of 24-h lighting and night feeding changed energy and carbohydrate metabolism to facilitate biosynthetic processes and, therefore, weight gain in fish.

Ginger Essential Oil Exerts Antibacterial Activity Against Shewanella putrefaciens by Disrupting Cell Structure and Resisting Biofilm

ABSTRACTThis study aimed to elucidate the antibacterial mechanism and antibiofilm activity of ginger essential oil (GEO) against Shewanella putrefaciens. On the basis of minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC), alkaline phosphatase (AKPase), succinate dehydrogenase (SDH), and glucose‐6‐phosphate dehydrogenase (G6PDH) activities were analyzed to evaluate the damage degree of S. putrefaciens on cell wall and membrane, and the microstructure was observed by scanning electron microscope (SEM) to evaluate the action effect against S. putrefaciens. The results showed that the MIC and MBC of GEO against S. putrefaciens were 27.8125 mg/mL and 55.625 mg/mL, respectively. The surface structures and cell membrane of S. putrefaciens were rigorously damaged by 1 MIC and 2 MIC GEO, leading to the leakage of cellular nucleic acids, protein, and β‐galactosidases from the bacterial cells. GEO could not only decrease the biomass of biofilm but also remove mature biofilm. In addition, GEO could reduce the production of exopolysaccharides and extracellular proteins, and could lessen the metabolic activity of biofilm cells. These results demonstrated that GEO exhibited efficient antibacterial characteristics against S. putrefaciens, which could act as a natural antibacterial and antibiofilm agent on the preservation of aquatic products.

Increased exercise tolerance in humanized G6PD deficient mice

Glucose-6-phosphate dehydrogenase (G6PD) deficiency affects 500 million people globally, impacting red blood cell (RBC) antioxidant pathways and increasing susceptibility to hemolysis under oxidative stress. Despite the systemic generation of reactive oxygen species during exercise, the effects of exercise on individuals with G6PD deficiency remain poorly understood This study utilized humanized mouse models expressing the G6PD Mediterranean variant (S188F, with 10% enzymatic activity) to investigate exercise performance and molecular outcomes. Surprisingly, despite decreased enzyme activity, G6PD-deficient mice have faster critical speed (CS) compared to mice expressing human canonical G6PD. Post-exercise, deficient mice did not exhibit differences in RBC morphology or hemolysis, but had improved cardiac function, including cardiac output, stroke volume, sarcomere length and mitochondrial content. Proteomics analyses of cardiac and skeletal muscles (gastrocnemius, soleus) from G6PD deficient compared to sufficient mice revealed improvements in mitochondrial function and increased protein turnover via ubiquitination, especially for mitochondrial and structural myofibrillar proteins. Mass spectrometry-based metabolomics revealed alterations in energy metabolism and fatty acid oxidation. These findings challenge the traditional assumptions regarding hemolytic risk during exercise in G6PD deficiency, suggesting a potential metabolic advantage in exercise performance for individuals carrying non-canonical G6PD variants.

Prevalence and screening of hemoglobinopathies and glucose-6-phosphate dehydrogenase deficiency in Yemeni blood donors

ABSTRACT Hemoglobinopathies and Glucose-6-Phosphate Dehydrogenase (G6PD) deficiency pose significant health threats in Yemen, which can be alleviated through robust preventive strategies, highlighting the importance of implementing a cost-effective and efficient blood donor screening method to facilitate early diagnosis and management of erythrocyte disorders, ultimately improving both donor safety and patient health outcomes. Objectives This study aimed to determine the prevalence of total blood cell abnormalities, hemoglobinopathies and G6PD deficiency and evaluate the efficacy of red blood cell (RBC) indices, mentzer index (MI) and naked-eye single tube red cell osmotic fragility (NESTROF) test as screening tools for diagnosis of β thalassemia trait among Yemeni blood donors. Methods A cross-sectional study was conducted with 106 volunteer blood donors who met the national standard criterion of blood donation. Various tests were performed, including complete blood count (CBC), serum ferritin, sickling test, G6PD assay, NESTROF test and high-performance liquid chromatography (HPLC). Results The prevalence of hematological abnormalities among blood donors reached 68.9%, with functional RBC abnormalities at 51.9%, leukopenia at 10.4%, thrombocytosis at 1.9%, and thrombocytopenia at 4.7%. Additionally, hemoglobinopathies were found in 21.7% of donors, with β-thalassemia trait at 3.8%, sickle cell trait at 1.9%, and suspected α-thalassemia trait at 16%, while G6PD deficiency and iron deficiency were present in 12.3% and 17.9% of donors, respectively. The NESTROF test, MCV and MCH demonstrated a sensitivity rate of 100%. MI and MCH exhibited the highest specificity followed by NESTROF test in the screening of β-thalassemia trait. Conclusions The prevalence of hemoglobinopathies and G6PD deficiency appear to be common among Yemeni blood donors. These results emphasize the necessity of comprehensive blood donation screening programs to safeguard the blood supply and promote early detection and management of hemoglobinopathies and G6PD deficiency in Yemen.

Optimization of a high throughput screening platform to identify inhibitors of asymmetric diadenosine polyphosphatases

When stressed, cells synthesize di-adenosine polyphosphates (ApnA), and cellular organisms also express proteins that degrade these compounds to release ATP. Most of these proteins are members of the nudix hydrolase superfamily, and several are involved in bacterial pathogenesis, neurodevelopment, and cancer. The goal of this project is to assist in the discovery of inhibitors of these enzymes that could be used to study ApnA function and the cellular role of these nudix enzymes. Because these enzymes cleave Ap4A and Ap5A to produce ATP, two standard ATP detection techniques were optimized and compared here for their suitability for high throughput screening. In the first assay, cleavage is monitored by coupling to a reaction catalyzed by firefly luciferase. In the second assay, cleavage is detected by coupling to hexokinase, glucose 6-phosphate dehydrogenase, and diaphorase. Although the former assay was more sensitive, the latter was more reproducible, linear, and suitable for screening and kinetic analyses. The assays were used to characterize the kinetics of reactions catalyzed by various nudix enzymes isolated from E. coli, humans, and Mycobacterium tuberculosis, the bacterium that causes tuberculosis. Results reveal subtle differences between the proteins that might be exploited to identify specific small molecule inhibitors.

Engineered Cardiac Tissues as a Platform for CRISPR-Based Mitogen Discovery.

Improved understanding of cardiomyocyte (CM) cell cycle regulation may allow researchers to stimulate pro-regenerative effects in injured hearts or promote maturation of human stem cell-derived CMs. Gene therapies, in particular, hold promise to induce controlled proliferation of endogenous or transplanted CMs via transient activation of mitogenic processes. Methods to identify and characterize candidate cardiac mitogens in vitro can accelerate translational efforts and contribute to the understanding of the complex regulatory landscape of CM proliferation and postnatal maturation. In this study, A CRISPR knockout-based screening strategy using in vitro neonatal rat ventricular myocyte (NRVM) monolayers is established, followed by candidate mitogen validation in mature 3-D engineered cardiac tissues (ECTs). This screen identified knockout of the purine metabolism enzyme adenosine deaminase (ADA-KO) as an effective pro-mitogenic stimulus. RNA-sequencing of ECTs further reveals increased pentose phosphate pathway (PPP) activity as the primary driver of ADA-KO-induced CM cycling. Inhibition of the pathway's rate limiting enzyme, glucose-6-phosphate dehydrogenase (G6PD), prevented ADA-KO induced CM cycling, while increasing PPPactivityviaG6PD overexpression increased CM cycling. Together, this study demonstrates the development and application of a genetic/tissue engineering platform forin vitrodiscovery and validation of new candidate mitogens affecting regenerative or maturation states of cardiomyocytes.

Identification of imidazo[1,2-a]pyridines-pyrazole-pyran hybrids as potent glucose-6-phosphate dehydrogenase (G6PD) inhibitors: Synthesis, molecular docking, DFT studies and ADME prediction

In this study, we aimed to identify new anticancer agents by synthesizing a series of Imidazo[1,2-a]pyridines fused with pyrazole-pyran scaffolds 7a–g in good yields (75–85 %) under microwave irradiation. The synthesis was performed through a one-pot, three-component cyclocondensation reaction of 2-(p-substituted)imidazo[1,2-a]pyridine-3-carbaldehyde 4a-g, malononitrile 5, and 5-methyl-2,4-dihydro-3H-pyrazol-3-one 6 in the presence of triethylamine as a catalyst in acetonitrile. The characterization of all synthesized compounds was accomplished through various spectroscopic techniques. All the synthesized compounds were investigated for their anticancer potential in vitro, using an MTT-based assay against A549 lung cancer cell lines. Compounds 7b, 7e, and 7c demonstrated notable anti-proliferative activity against the A549 cell line, while compound 7e showed a significantly lower fluorescence generation (24-fold less) in the G6PD inhibition assay compared to the other compounds.

Phytochemical Analysis, Total Phenolic Content, Hemolytic and Anti-hemolytic Activities of Centaurea iberica (Asteraceae)

Several plants contain chemical substances that have various biological activities, but they also probably cause harmful effects on cellular membranes. This work deals with the Centaurea iberica (Iberian starthistle) plant. The objective of this study is to identify the main chemical groups in the methanolic extract 80% of the plant, determine its total phenolic content, evaluate its toxic effect on human erythrocytes, and also assess its activity in the protection of normal and glucose-6-phosphate dehydrogenase (G6PD) enzyme-deficient human erythrocytes membranes against oxidative hemolysis. Phytochemical analysis revealed that C. iberica extract contains several bioactive compounds. The results also showed that the plant extract contained good level of total phenolic compounds. The extract exhibited a low hemolytic effect. It also showed excellent activity in the protection of normal and G6PD-deficient human erythrocytes against oxidative hemolysis. KEYWORDS Erythrocytes, G6PD, Iberian starthistle, methanolic extract, toxic effect, oxidative hemolysis

Case report: Dichloroacetate-induced methaemoglobinaemia in a G6PD-deficient neonate.

A 3-week-old neonate with glucose-6-phosphate dehydrogenase (G6PD) deficiency and primary lactic acidosis developed haemolytic jaundice and methaemoglobinaemia following treatment with dichloroacetate (DCA), a standard treatment for primary lactic acidosis. While this mechanism has been reported in the sheep model, it has not been described in humans. Our case reinforces the uncommon observation that a G6PD-deficient individual experiencing oxidative stress may develop concurrent methaemoglobinaemia. In this case, methylene blue, the standard treatment for methaemoglobinaemia, may result in further oxidative stress. The judicious use of blood transfusion to correct the oxygen-carrying capacity of our patient led to reversal of the methaemoglobinaemia.

Molecular epidemiological characteristics, variant spectrum and genotype-phenotype correlation of glucose-6-phosphate dehydrogenase deficiency in China: A population-based multicenter study using newborn screening.

Newborn screening (NBS) for glucose-6-phosphate dehydrogenase (G6PD) deficiency by biochemical tests is being used worldwide, however, the outcomes arising from combined genetic and biochemical tests have not been evaluated. This research aimed to evaluate the outcomes of application of combined genetic and biochemical NBS for G6PD deficiency and to investigate the molecular epidemiological characteristics, variant spectrum, and genotype-phenotype correlation of G6PD deficiency in China. A population-based cohort of 29,601 newborns were prospectively recruited from eight NBS centers in China between February 21 and December 30, 2021. Biochemical and genetic NBS was conducted simultaneously. The overall prevalence of G6PD deficiency was 1.12% (1.86% for male, and 0.33% for female; 1.94% for South China and 0.08% for North China). Genetic NBS identified 10 male patients undetected by biochemical NBS. The overall positive predictive values (PPVs) of biochemical and genetic NBS were 79.95% and 47.57%, respectively. A total of 15 variants were identified, with the six most common variants being c.1388G > A, c.1376G > T, c.95A > G, c.871G > A, c.1024C > T and c.392G > T (94.2%). The activity of G6PD was correlated with the type and WHO classification of variants. This study highlighted that combined screening could enhance the efficiency of current NBS for diagnosing G6PD deficiency. The prevalence, variant spectrum and allele frequency of G6PD deficiency vary across different regions. Our data provide valuable references for clinical practice and optimization of future screening strategies for G6PD deficiency.

G>A transition at 376 position of the glucose-6-phosphate dehydrogenase (G6PD) gene plays a key role in causing G6PD deficiency in the Siddi population of Karnataka.

G>A transition at 376 position of the glucose-6-phosphate dehydrogenase (G6PD) gene plays a key role in causing G6PD deficiency in the Siddi population of Karnataka.

Supplemental oxygen alters the pentose phosphate pathway in the developing mouse brain through SIRT signaling

Oxygen support plays a critical role in the management of preterm infants in neonatal intensive care units. On the other hand, the possible effects of oxygen supplementation on cellular functions, specifically glucose metabolism, have been less understood. Purposeof the study is to investigate whether supplemental oxygen alters glucose metabolism and pentose phosphate pathway (PPP) activity in the brain tissue and its relevance with silent information regulator proteins (SIRT) pathway. For this purpose, newborn C57BL/6 pups were exposed to 90% oxygen from birth until postnatal day 7 (PN7) and metabolites of glysolysis and PPP were investigated through metabolomics analysis. SIRT1, glucose-6-phosphate dehydrogenase (G6PD) and transaldolase (TALDO) proteins were examined immunohistochemically and molecularly in the prefrontal and hippocampus regions of the brain. Later on, SIRT1 inhibition was carried out.Our results indicate that supplemental oxygen causes an increase in PPP metabolites as well as activation of G6PD enzyme in the brain tissue, which is reversed by SIRT1 inhibition. Our study underlines a connection between supplemental oxygen, glucose metabolism, PPP pathway and the SIRT signaling. Understanding these intricate relationships not only deepens our knowledge of cellular physiology but also holds promise for therapeutic interventions for creating neuroprotective strategies in preterm brain.

Expanding families: a pilot study on preconception expanded carrier screening in Bahrain

BackgroundPreconception expanded carrier screening (ECS) is a genetic test that enables the identification of at-risk carriers of recessive disorders by screening for up to hundreds of genes. Next-generation sequencing (NGS) development has paved the way for its integration into ECS. This study aims to identify the carrier genetic status of couples experiencing or anticipating conception challenges through NGS-based ECS and to gain an overview of the rare genetic disorders in a population with increased consanguinity.MethodsThirty couples who presented to the Genetic Disease Clinic between 2015 and 2024 with failed reproductive outcomes or with a positive personal or family history of genetic disorders and underwent ECS were included and retrospectively analyzed.ResultsFifty-four individuals (90.00%) were found to carry at least one variant of 95 identified genes, totaling 174 variants. Six individuals (10.00%) tested negative for any variant. Seven individuals had one variant (11.67%), 13 had two variants (21.67%), and 34 had 3 or more variants (56.67%). The most common variants identified were of HBA, HBB, CYP21A2, and G6PD genes. Most of the detected variants were unknown or unexpected (n = 143, 82.18%). Eight couples carried two or more variants in common. Consanguinity was reported in 14 couples (46.67%).ConclusionsPreconception ECS is crucial for reproductive planning, permitting couples to evaluate their combined genetic risks and make informed decisions, reducing the chance of having children with genetic disorders.

Buffy coat removal to mitigate reporting false normal G6PD phenotype results in patients undergoing rasburicase therapy for tumor lysis syndrome

Abstract Glucose-6-phosphate dehydrogenase (G6PD) is the primary source of NADPH in red blood cells (RBCs). NADPH reduces reactive oxygen species (ROS) thereby protecting RBCs from oxidative stress-mediated hemolytic anemia (HA). Individuals with partial or full G6PD deficiency are therefore susceptible to acute HA provoked by medications that generate high ROS loads in vivo, e.g., rasburicase. Rasburicase is an enzyme-based therapeutic used to degrade uric acid (UA) in patients with tumor lysis syndrome (TLS); however, catalysis of UA generates H2O2, a potent ROS, which poses a risk of HA in individuals who are deficient in G6PD activity. Screening G6PD activity prior to rasburicase administration is clinically indicated in ethnic groups with high rates of G6PD deficiency, although reports of HA in rasburicase-treated patients have been reported across low prevalence ethnic groups as well. TLS occurs most commonly during the treatment of highly proliferative neoplasms, such as leukemia and lymphoma. Since WBCs contain significant G6PD activity, an elevated WBC count in these patients can cause the false assurance of low HA risk due to a falsely normal G6PD activity. We sought to establish and evaluate a novel method to mitigate WBC contamination of G6PD measurements in patients undergoing rasburicase therapy. In this study, we first developed and validated an innovative method (“piercing method”) to minimize the interference of WBCs in G6PD measurements. The piercing method demonstrated acceptable accuracy (slope: 1.008; bias: -6.22%) and precision (1.4% CV between 9.9 to 20.7 U/g Hb) and was effective at eliminating a broad range of WBC types and concentrations from whole blood samples (N = 85, tested range = 0.31 – 413.05 K/μL, 93% reduction in WBCs). We compared G6PD activity in rasburicase-treated patients (N = 39) with varying WBC counts before and after buffy coat removal using the piercing method. Average percent change in G6PD activity (post-pierce – pre-pierce) between patients with and without elevated WBCs was substantial (45% versus 12%). In comparing G6PD activity in patients before and after the initiation of rasburicase therapy, there is no evidence that in vivo-generated ROS from rasburicase affect G6PD measurement in vitro in an assay that relies on monitoring absorbance of NADPH (22.2 U/g Hb versus 22.9 U/g Hb, p=0.721). Results demonstrate how WBCs increase risk for false elevation in G6PD activity in patients being considered for rasburicase therapy. At greatest risk are patients with partial G6PD deficiency, e.g., heterozygous females, where G6PD from WBCs could mask deficiency. Literature describing testing of G6PD in specimens with elevated WBCs is lacking. This study shows that the piercing method provides accurate G6PD results in samples with elevated WBCs from patients at risk of TLS, who might develop HA after starting rasburicase therapy.

Change in Activities of Enzymes of Energy and Carbohydrate Metabolism in Pink Salmon Oncorhynchus gorbuscha (Walb.) Smolts with Change in Environmental Salinity.

Activities of key enzymes of energy and carbohydrate metabolism (cytochrome c oxidase (COX), lactate dehydrogenase (LDH), aldolase, glucose 6-phosphate dehydrogenase (G6PDH), and 1-glycerophosphate dehydrogenase (1-GPDH)) were studied in pink salmon Oncorhynchus gorbuscha (Walb.) smolts from the White Sea in a cage experiment simulating the transition from a freshwater to a marine environment. A decrease in COX, G6PDH, and 1-GPDH activities and an increase in LDH and aldolase activities were observed in juveniles with an increase in water salinity. Based on the findings, a redistribution of energy substrates between the reactions of aerobic and anaerobic metabolism towards higher anaerobic ATP synthesis was assumed for pink salmon. This may indicate that adaptive mechanisms rearrange metabolism to provide energy for osmoregulation in pink salmon juveniles when the salinity changes in their habitat.

Identification of a novel NADPH generation reaction in the pentose phosphate pathway in Escherichia coli using mBFP.

NADPH is a redox cofactor that drives the anabolic reactions. Although major NADPH generation reactions have been identified in Escherichia coli, some minor reactions have not been identified. In the present study, we explored novel NADPH generation reactions by monitoring the fluorescence dynamics after the addition of carbon sources to starved cells, using a metagenome-derived blue fluorescent protein (mBFP) as an intracellular NADPH reporter. Perturbation analyses were performed on a glucose-6-phosphate isomerase (PGI) deletion strain and its parental strain. Interestingly, mBFP fluorescence increased not only in the parental strain but also in the ΔPGI strain after the addition of xylose. Because the ΔPGI strain cannot metabolize xylose through the oxidative pentose phosphate pathway, this suggests that an unexpected NADPH generation reaction contributes to an increase in fluorescence. To unravel this mystery, we deleted the NADPH generation enzymes including transhydrogenase, isocitrate dehydrogenase, NADP+-dependent malic enzyme, glucose-6-phosphate dehydrogenase (G6PDH), and 6-phosphogluconate dehydrogenase (6PGDH) in the ΔPGI strain, and revealed that G6PDH and 6PGDH contribute to an increase in fluorescence under xylose conditions. In vitro assays using purified enzymes showed that G6PDH can produce NADPH using erythrose-4-phosphate (E4P) as a substitute for glucose-6-phosphate. Because the Km (0.65 mM) for E4P was much higher than the reported intracellular E4P concentrations in E. coli, little E4P must be metabolized through this bypass in the parental strain. However, the flux would increase when E4P accumulates in the cells owing to genetic modifications. This finding provides a metabolic engineering strategy for generating NADPH to produce useful compounds using xylose as a carbon source.IMPORTANCEBecause NADPH is consumed during the synthesis of various useful compounds, enhancing NADPH regeneration is highly desirable in metabolic engineering. In this study, we explored novel NADPH generation reactions in Escherichia coli using a fluorescent NADPH reporter and found that glucose-6-phosphate dehydrogenase can produce NADPH using erythrose-4-phosphate as a substrate under xylose conditions. Xylose is an abundant sugar in nature and is an attractive carbon source for bioproduction. Therefore, this finding contributes to novel pathway engineering strategies using a xylose carbon source in E. coli to produce useful compounds that consume NADPH for their synthesis.

Stoichiometric Regeneration of Biomimetic Nicotinamide Coenzyme Powered by Biomass Sugars via In Vitro Synthetic Enzymatic Biosystems.

Biomimetic nicotinamide coenzymes, including nicotinamide mononucleotide (NMN+), have been demonstrated as promising low-cost alternatives to nicotinamide adenine dinucleotide (phosphate) (NAD(P)+) in biocatalysis. Herein, to efficiently regenerate NMNH from NMN+ in vitro powered by biomass sugars, a thermophilic NADP+-dependent glucose 6-phosphate dehydrogenase from Thermotoga maritima (TmG6PDH) was engineered to increase the activity toward NMN+. The catalytic efficiency (kcat/Km) of optimal mutant (TmG6PDH-R7) toward NMN+ increased by 71.7-fold than TmG6PDH-WT. As a result, compared to the wild type, the coenzyme specificity ([kcat/Km]NMN+/[kcat/Km]NADP+) of TmG6PDH-R7 increased by ~2.0×105-fold. The structural analysis revealed that the introduced hydrophobic and bulky residues lead to the formation of a smaller binding pocket, which resulting in a higher affinity for NMN+ with small size than NADP+. Then several in vitro synthetic enzymatic biosystems (ivSEBs) comprising this thermophilic TmG6PDH-R7 and a previously engineered thermophilic 6-phosphogluconate dehydrogenase were constructed. These ivSEBs harnessed the complete oxidation of renewable biomass sugars to facilitate the stoichiometric regeneration of 12 molecules of NMNH from 1 molecule of glucose, thereafter producing various products such as levodione, 2,3-butanediol or bioelectricity, over a wide temperature range. This study could pave the way for using stable and low-cost biomimetic coenzymes in ivSEBs for industrial biomanufacturing.

In Silico Approach for Assessment of the Anti-Tumor Potential of Cannabinoid Compounds by Targeting Glucose-6-Phosphate Dehydrogenase Enzyme.

Glucose-6-phosphate dehydrogenase (G6PD) is a pentose phosphate pathway (PPP) enzyme that generates NADPH, which is required for cellular redox equilibrium and reductive biosynthesis. It has been demonstrated that abnormal G6PD activation promotes cancer cell proliferation and metastasis. To date, no G6PD inhibitor has passed clinical testing successfully enough to be launched as a medicine. As a result, in this investigation, cannabinoids were chosen to evaluate their anticancer potential by targeting G6PD. Molecular docking indicated that three molecules, Tetrahydrocannabinolic acid (THCA), Cannabichromenic acid (CBCA), and tetrahydrocannabivarin (THCV), have the highest binding affinities for G6PD of -8.61, -8.39, and 8.01 Kcal mol. ADMET analysis found that all of them were safe prospective drug candidates. Molecular dynamics (MD) simulation and MM-PBSA analysis confirm the structural compactness and lower conformational variation of protein-ligand complexes, thereby maintaining structural stability and rigidity. Thus, our in silico investigation exhibited all three cannabinoids as potential competitive inhibitors of G6PD.

Bioorthogonal Regulated Metabolic Balance for Promotion of Ferroptosis and Mild Photothermal Therapy.

The nanozyme with NADPH oxidase (NOX)-like activity can promote the consumption of NADPH and the generation of free radicals. In consideration of that the upregulation of glucose-6-phosphate dehydrogenase (G6PD) would accelerate the compensation production of NADPH, for inhibition of G6PD activity, our designed bioorthogonal nanozyme can in situ catalyze pro-DHEA to produce G6PD inhibitor and dehydroepiandrosterone (DHEA) drugs to inhibit G6PD activity. Therefore, the well-defined platform can disrupt NADPH homeostasis, leading to the collapse of the antioxidant defense system in the tumor cells. The enzyme-like activity of PdCuFe is further enhanced when irradiated by NIR-II light. The destruction of NADPH homeostasis can promote ferroptosis and, in turn, facilitate mild photothermal therapy. Our design can realize NADPH depletion and greatly improve the therapeutic effect through metabolic regulation, which may provide inspiration for the design of bioorthogonal catalysis.