Enzymes In Metabolic Pathways Research Articles

Pharmacotherapeutic Potential of Bitter Gourd (Momordica charantia) in Age-related Neurological Diseases.

Due to the growth of the elderly population, age-related neurological disorders are an increasing problem. Aging begins very gradually and later leads to several neurological issues such as lower neurotransmitter levels, oxidative stress, neuronal inflammation, and continual neuronal loss. These changes might contribute to brain disorders such as Alzheimer's disease (AD), dementia or mild cognitive impairment, and epilepsy and glioma, and can also aggravate these disorders if they were previously present. Momordica charantia (bitter gourd), a member of the Cucurbitaceae family, is a good source of carbohydrates, proteins, vitamins, and minerals. It is used for diabetes and known for its hypoglycemic and antioxidant effects. In this review, we discuss the pharmaceutical effects of M. charantia on age-related neurological disorders. We searched several databases, including PubMed and Google Scholar, using MeSH terms. We searched articles published up until 2022 regardless of publication language. M. charantia is rich in luteolin, which increases acetylcholine in neurons by binding to enzymes in acetylcholine metabolism pathways, including butyrylcholinesterase and acetylcholinesterase. This binding inhibits the hyperphosphorylation of tau protein by restraining its kinase enzyme. Furthermore, this substance can lower serum cholesterol and has multi-target activity in AD and memory loss. M. charantia can also improve memory by decreasing tau protein and it also has potent antioxidant activity and anti-inflammatory effects. This review highlights that M. charantia has effects on many age-related neurological disorders, and can be a cost-effective supplement with minimal side effects.

Rapid HPLC method reveals dynamic shifts in coenzyme Q redox state

Ubiquinol or coenzyme Q (CoQ) is a lipid-soluble electron carrier in the respiratory chain and an electron acceptor for various enzymes in metabolic pathways that intersect at this cofactor hub in the mitochondrial inner membrane. The reduced form of CoQ is an antioxidant, which protects against lipid peroxidation. In this study, we have optimized a UV-detected HPLC method for CoQ analysis from biological materials, which involves a rapid single-step extraction into n-propanol followed by direct sample injection onto a column. Using this method, we have measured the oxidized, reduced and total CoQ pools, and monitored shifts in the CoQ redox status in response to cell culture conditions and bioenergetic perturbations. We find that hypoxia or sulfide exposure induces a reductive shift in the intracellular CoQ pool. The effect of hypoxia is however, rapidly reversed by exposure to ambient air. Interventions at different loci in the electron transport chain can induce sizeable redox shifts in the oxidative or reductive direction, depending on whether they are up- or downstream of complex III. We have also used this method to confirm that CoQ levels are higher and more reduced in murine heart versus brain. In summary, the availability of a convenient HPLC-based method described herein, will facilitate studies on CoQ redox dynamics in response to environmental, nutritional and endogenous alterations.

Unveiling the Biological Function of Phyllostachys edulis FBA6 (PeFBA6) through the Identification of the Fructose-1,6-Bisphosphate Aldolase Gene.

Fructose-1,6-bisphosphate aldolase (FBA) is a pivotal enzyme in various metabolic pathways, including glycolysis, gluconeogenesis, and the Calvin cycle. It plays a critical role in CO2 fixation. Building on previous studies on the FBA gene family in Moso bamboo, our study revealed the biological function of PeFBA6. To identify CSN5 candidate genes, this study conducted a yeast two-hybrid library screening experiment. Subsequently, the interaction between CSN5 and PeFBA6 was verified using yeast two-hybrid and LCI experiments. This investigation uncovered evidence that FBA may undergo deubiquitination to maintain glycolytic stability. To further assess the function of PeFBA6, it was overexpressed in rice. Various parameters were determined, including the light response curve, CO2 response curve, and the levels of glucose, fructose, sucrose, and starch in the leaves of overexpressing rice. The results demonstrated that overexpressed rice exhibited a higher saturation light intensity, net photosynthetic rate, maximum carboxylation rate, respiration rate, and increased levels of glucose, fructose, and starch than wild-type rice. These findings indicated that PeFBA6 not only enhanced the photoprotection ability of rice but also improved the photosynthetic carbon metabolism. Overall, this study enhanced our understanding of the function of FBA and revealed the biological function of PeFBA6, thereby providing a foundation for the development of excellent carbon fixation bamboo varieties through breeding.

Photoheterotroph improved the growth and nutrient levels of Chlorella vulgaris and the related molecular mechanism

Microalgae are rich in fatty acids, proteins, and other nutrients, which have gained the general attention of researchers all over the world. For the development of Chlorella vulgaris in food and feed industry, this study was conducted to investigate the differences in C. vulgaris’ growth and nutritional components under different culture conditions (autotrophic, heterotrophic, photoheterotrophic) and the internal factors through cell counting in combination with transcriptome and nutrient analyses. The results showed that, under the photoheterotrophic condition, Chlorella’s growth and the contents of lipid and protein were significantly higher than that under the heterotrophic condition, and the moisture content was lower than that under the heterotrophic condition. The saturated fatty acid content under the photoheterotrophic condition was the lowest, while the polyunsaturated fatty acid content was significantly higher than those under the other two conditions. There were 46,583 differentially expressed genes (DEGs), including 33,039 up-regulated DEGs (70.93%) and 13,544 down-regulated DEGs (29.07%), under the photoheterotrophic condition in comparison with the autotrophic condition. The fold change between the two conditions of samples of up-regulated genes was higher than that of the down-regulated genes. The KEGG enrichment showed that the up-regulated DEGs in the photoheterotrophic condition were significantly enriched in 5 pathways, including protein processing in endoplasmic reticulum pathway, photosynthesis pathway, photosynthesis-antenna protein pathway, endocytosis pathway, and phosphonate and phosphinate metabolism pathway. DEGs related to fatty acid metabolic pathways were significantly enriched in the fatty acid biosynthesis pathway and the biosynthesis of unsaturated fatty acid pathway. The qPCR analysis showed that the expression pattern of the selected genes was consistent with that of transcriptome analysis. The results of this study lay a theoretical foundation for the large-scale production of Chlorella and its application in food, feed, and biodiesel.Key points• Nutrient levels under photoheterotrophic condition were higher than other conditions.• Six important pathways were discovered that affect changes in nutritional composition.• Explored genes encode important enzymes in the differential metabolic pathways.

Senescence and aging differentially alter key metabolic pathways in murine brain microglia

Microglia, the resident immune cells of the central nervous system, are critically involved in maintaining brain homeostasis. With age, microglia display morphological and functional alterations that have been associated with cognitive decline and neurodegeneration. Although microglia seem to participate in an increasing number of biological processes which require a high energy demand, little is known about their metabolic regulation under physiological and pathophysiological conditions and during aging/senescence. Here, we determined mRNA expression levels of critical rate limiting enzymes in several key metabolic pathways including glycolysis, pentose phosphate pathway, fatty acid oxidation and synthesis in association with oxidative phosphorylation in microglia, both under aging and senescent conditions. We found strong evidence for different metabolic changes occuring in senescent vs. aged microglia cells. While senescent microglia display a hypermetabolic state as indicated by increased expression of key enzymes involved in glycolysis and pentose phosphate pathway, aging microglia are rather in a state of hypometabolism. Our findings indicate that studies involving aging and senescent microglia require a clear differentiation between these microglial states due to profound metabolic differences observed here. Understanding metabolic changes in senescent and aged microglia may lead to novel strategies to decrease over-activation of these cells due to aging, which is associated to the process of inflamm-aging and neurodegeneration.

Enhanced branched-chain amino acid metabolism improves age-related reproduction in C. elegans.

Reproductive ageing is one of the earliest human ageing phenotypes, and mitochondrial dysfunction has been linked to oocyte quality decline; however, it is not known which mitochondrial metabolic processes are critical for oocyte quality maintenance with age. To understand how mitochondrial processes contribute to Caenorhabditis elegans oocyte quality, we characterized the mitochondrial proteomes of young and aged wild-type and long-reproductive daf-2 mutants. Here we show that the mitochondrial proteomic profiles of young wild-type and daf-2 worms are similar and share upregulation of branched-chain amino acid (BCAA) metabolism pathway enzymes. Reduction of the BCAA catabolism enzyme BCAT-1 shortens reproduction, elevates mitochondrial reactive oxygen species levels, and shifts mitochondrial localization. Moreover, bcat-1 knockdown decreases oocyte quality in daf-2 worms and reduces reproductive capability, indicating the role of this pathway in the maintenance of oocyte quality with age. Notably, oocyte quality deterioration can be delayed, and reproduction can be extended in wild-type animals both by bcat-1 overexpression and by supplementing with vitamin B1, a cofactor needed for BCAA metabolism.

The impact of some metals, molecular docking and molecular dynamic calculations on glucose 6-phosphate dehydrogenase activity in Capoeta trutta (Heckel, 1843) tissue

The first enzyme of the pentose phosphate metabolic pathway is glucose 6-phosphate dehydrogenase (d-glucose-6-phosphate: NADP + oxidoreductase EC1.1.1.49; G6PD). G6PD has essential functions such as membrane lipid synthesis, ribose 5-phosphate, and NADPH production. In this study, the G6PD enzyme was purified from kidney, liver, and gill tissues of Capoeta trutta, one of the dominant fish species in the Euphrates-Tigris River System, and the in vitro effects of some metals (Ag+, Cd2+, Cu2+, Fe2+, Ni2+, Pb2+ and Zn2+) on the enzyme activity were investigated. For this purpose, firstly, the G6PD enzyme was purified from tissues using a 2′, 5′-ADP Sepharose 4B affinity column. The purity of the enzyme was checked by the SDS-PAGE method and a single band was seen in the gel. After the purity of the enzyme was determined, the effects of metals on the enzyme activity were determined using the spectrophotometric method. As a result of the study, it was determined that the Ag+ ion was the most potent inhibitor for C. trutta gill, kidney, and liver G6PD enzymes. Lastly, calculations were made to examine the activity of the glucose 6-phosphate dehydrogenase molecule against the G6PD enzymes. Afterwards, the interaction of the glucose 6-phosphate dehydrogenase molecule with the protein (PDB ID: 5JYU and 2BH9) with the highest activity was calculated in the range of 0–100 ns. Finally, ADME/T calculation was made to predict the effects and reactions of glucose 6-phosphate dehydrogenase molecule in human metabolism. This study explores the physiological functions and environmental sensitivities of G6PD in a dominant fish species, while also investigating its potential interactions and metabolic roles in humans. Understanding these aspects can contribute to environmental monitoring, fish health management, and even pharmaceutical development.

Functional identification of AeHMGR gene involved in regulation of saponin biosynthesis in Aralia elata

Aralia elata (Miq.) Seem, a significant tree species in the Araliaceae family, has medicinal and edible properties. Saponins are the primary active components of A. elata. The 3-hydroxy-3-methylglutaryl- CoA reductase (HMGR) is the initial rate-limiting enzyme of the major metabolic pathway of saponins in A. elata. In this study, the AeHMGR gene was identified through screening of transcriptome data. Through the qRT-PCR analysis, it was determined that the expression level of AeHMGR gene is highest in the somatic embryo and stem of A. elata. Heterologous transformation in tobacco revealed that ectopic expression of the AeHMGR gene leads to a significant reduction in the expression levels of the NtSS, NtFPS, and NtSE genes in transgenic tobacco lines, with a minimum expression level of 0.24 times that of the wild type. In the overexpressed callus lines of A. elata, the expression levels of the AeFPS, AeSE, AeSS, and Aeβ-AS genes were also significantly lower compared to the wild type, with a minimum expression level of approximately 0.3 times that of the wild type. Interestingly, the overexpression of the AeHMGR gene in A. elata somatic embryos led to a substantial decrease in the expression levels of AeFPS and AeSS, while the expression levels of AeSE and Aeβ-AS increased. Among the transgenic somatic embryo strain lines, line 7 exhibited the highest expression levels of AeSE and Aeβ-AS, with fold increases of 11.51 and 9.38, respectively, compared with that of the wild-type. Additionally, a high-performance liquid chromatography method was established to detect five individual saponins in transgenic A. elata. The total saponin content in line 7 somatic embryos was 1.14 times higher than that of wild-type materials, but only 0.30 times that of wild-type cultivated leaves. Moreover, the content of oleanolic acid saponin in line 7 was 1.35 times higher than that of wild-type cultivated leaves. These indicate that HMGR can affect triterpene biosynthesis.

A major facilitator superfamily transporter MdtH in Escherichia coli is involved in anthocyanin biosynthesis and secretion.

Cyanidin 3-O-glucoside (C3G) is a natural colorant with health-promoting activities and is, hence, widely used in food, cosmetic, and nutraceutical industries. Its market supply is currently dependent on extraction from plants. As an alternative, C3G can be produced by the microbe Escherichia coli in a green and sustainable way. However, a large portion of this compound is retained inside the cell of E. coli, thus complicating the purification process and limiting the high-level production. We have identified and verified an efficient native transporter named MdtH in E. coli that can export C3G to the cultivation medium. Overexpression of MdtH could improve extracellular C3G production by 110% without modifications of the metabolic pathway genes or enzymes. This study reveals a new regulation target for C3G production in bacteria and provides guidance to the microbial biosynthesis of related compounds.

Optimal enzyme profiles in unbranched metabolic pathways.

How to optimize the allocation of enzymes in metabolic pathways has been a topic of study for many decades. Although the general problem is complex and nonlinear, we have previously shown that it can be solved by convex optimization. In this paper, we focus on unbranched metabolic pathways with simplified enzymatic rate laws and derive analytic solutions to the optimization problem. We revisit existing solutions based on the limit of mass-action rate laws and present new solutions for other rate laws. Furthermore, we revisit a known relationship between flux control coefficients and enzyme abundances in optimal metabolic states. We generalize this relationship to models with density constraints on enzymes and metabolites, and present a new local relationship between optimal reaction elasticities and enzyme amounts. Finally, we apply our theory to derive simple kinetics-based formulae for protein allocation during bacterial growth.

Co-metabolism driven sulfaquinoxaline removal in microbial electrolysis cells: A mechanistic analysis based on DFT calculation, metabolic pathway and functional enzyme activity

Sulfaquinoxaline (SQX) is a vital sulfonamide antibiotic for the treatment of livestock infections. Although its degradation has become a research focus in the last few years, its biodegradation process and biometabolic mechanisms were rarely studied. In this research, microbial electrolytic cells (MECs) were used to remove SQX using sodium acetate as a co-substrate. The findings indicated that the maximum removal rate of SQX for a 5-day period in co-substrate reached 94.2%, and SQX exhibited a lower resistance (45.64 Ω) and had a larger double-layer area than a single substrate (95.03 Ω). LCMS/MS analysis suggested that SQX could eventually degrade to 17 byproducts through hydrolysis, hydroxylation, N rearrangement and sulfur reduction. Density flooding theory (DFT) calculations showed that sodium acetate addition weakened the N-C bond and facilitated hydroxylation. Meanwhile, the fluorescence composition of fDOM shifted from protein-like to humic acid-like with co-substrate supplementation. Microbial community analysis showed that Actinobacteria, Proteobacteria, Bacteroidota and Chloroflexi were highly correlated with SQX removal. Analysis of the metabolic pathways showed that key enzymes such as NAT, CAT, DHP and NAD+ were involved in the degradation of SQX. Biotoxicity tests indicated that weak electrical stimulation was beneficial in reducing biotoxicity. These results provide theoretical basis and new insights into the bioelectrochemical degradation of SQX.

Mechanism of inactivation of Aspergillus flavus spores by dielectric barrier discharge plasma

Dielectric barrier discharge plasma (DBDP) displays strong against fungal spores, while its precise mechanism of spore inactivation remains inadequately understood. In this study, we applied morphological, in vivo and in vitro experiments, transcriptomics, and physicochemical detection to unveil the potential molecular pathways underlying the inactivation of Aspergillus flavus spores by DBDP. Our findings suggested that mycelium growth was inhibited as observed by SEM after 30 s treatment at 70 kV, meanwhile spore germination ceased and clustering occurred. It led to the release of cellular contents and subsequent spore demise by disrupting the integrity of spore membrane. Additionally, based on the transcriptomic data, we hypothesized that the induction of spore inactivation by DBDP might be associated with downregulation of genes related to cell membranes, organelles (mitochondria), oxidative phosphorylation, and the tricarboxylic acid cycle. Subsequently, we validated our transcriptomic findings by measuring the levels of relevant enzymes in metabolic pathways, such as superoxide dismutase, acetyl-CoA, total dehydrogenase, and ATP. These physicochemical indicators revealed that DBDP treatment resulted in mitochondrial dysfunction, redox imbalance, and inhibited energy metabolism pathways. These findings were consistent with the transcriptomic results. Hence, we concluded that DBDP accelerated spore rupture and death via ROS-mediated mitochondrial dysfunction, which does not depend on cell membranes.

Fused Enzyme Glucose-6-Phosphate Dehydrogenase::6-Phosphogluconolactonase (G6PD::6PGL) as a Potential Drug Target in Giardia lamblia, Trichomonas vaginalis, and Plasmodium falciparum.

Several microaerophilic parasites such as Giardia lamblia, Trichomonas vaginalis, and Plasmodium falciparum are major disease-causing organisms and are responsible for spreading infections worldwide. Despite significant progress made in understanding the metabolism and molecular biology of microaerophilic parasites, chemotherapeutic treatment to control it has seen limited progress. A current proposed strategy for drug discovery against parasitic diseases is the identification of essential key enzymes of metabolic pathways associated with the parasite's survival. In these organisms, glucose-6-phosphate dehydrogenase::6-phosphogluconolactonase (G6PD:: 6PGL), the first enzyme of the pentose phosphate pathway (PPP), is essential for its metabolism. Since G6PD:: 6PGL provides substrates for nucleotides synthesis and NADPH as a source of reducing equivalents, it could be considered an anti-parasite drug target. This review analyzes the anaerobic energy metabolism of G. lamblia, T. vaginalis, and P. falciparum, with a focus on glucose metabolism through the pentose phosphate pathway and the significance of the fused G6PD:: 6PGL enzyme as a therapeutic target in the search for new drugs.

Differential effects of heart failure with reduced ejection fraction on locomotor and respiratory muscles

Abstract Background Muscle weakness impairs quality-of-life, exercise capacity, and survival in patients with heart failure and reduced left ventricular ejection fraction (HFrEF). However, the effects on systemic concentrations and local expression of inflammatory cytokines and myokines as well as metabolic markers might differ between locomotor and respiratory muscles. Purpose We prospectively evaluated clinical, functional, and molecular alterations of the diaphragm and the M. vastus lateralis (MVL) from HFrEF patients receiving a left ventricular assist device compared to patients without heart failure undergoing elective coronary bypass grafting (control). Methods Participants (Controls=21, HFrEF=18) underwent cardiopulmonary exercise and respiratory and leg muscle testing. Serum concentrations and local expression of inflammatory cytokines and myokines as well as key enzymes of metabolic pathways were examined in biopsies of the diaphragm and MVL. Results Exercise capacity, respiratory muscle function and endurance force of locomotor muscles were significantly reduced in HFrEF compared with controls (all p<0.05). The serum concentration of Il-6, Il-1beta and TNF-alpha were significantly higher or solely detectable in HFrEF compared with controls. Local mRNA expression of Il-6 was not different in MVL but 2.2-fold higher in the diaphragm of HFrEF compared with controls (p=0.013). In contrast, mRNA expression of Il-6 receptor was significantly reduced by 38% (p=0.004) in MVL of HFrEF compared with controls but comparable in the diaphragm. Serum concentrations of FGF-21 and Musclin were significantly higher in HFrEF compared with controls (all p<0.05). Local mRNA expression of FGF-21 was significantly lower in both MVL and diaphragm of HFrEF compared with controls. FGF-21 receptor 1 mRNA expression was reduced by 18% in the MVL of HFrEF (p=0.02), whereas co-receptor klotho was comparable to controls. In the diaphragm, FGF-21 receptor 1 mRNA expression was comparable between HFrEF and controls, but the co-receptor klotho had a 5-fold higher mRNA expression in HFrEF (p=0.002). We examined the activity of key enzymes of glucose, lipid and ketone metabolism as well as key enzymes of the Krebs cycle. The main differences were found for ß-hydroxyacyl-CoA Dehydrogenase in the diaphragm with a 1.6-fold higher activity in HFrEF compared with controls (p=0.01) and for succinyl-CoA: 3-ketoacid-coenzyme A transferase in the MVL with a 2-fold higher activity in HFrEF compared with controls (p<0.01) indicating a main utilization of lipids in the diaphragm and of ketones in the MVL for energy supply. Conclusions HFrEF induced divergent effects on inflammatory markers, myokine expression and metabolic parameters in locomotor and respiratory muscles in HFrEF patients compared with controls. This indicates that different molecular alterations contribute to locomotor/respiratory muscle dysfunction and reduced whole-body exercise capacity.

PMPTCE-HNEA: Predicting Metabolic Pathway Types of Chemicals and Enzymes with a Heterogeneous Network Embedding Algorithm

Background: Metabolic chemical reaction is one of the main types of fundamental processes to maintain life. Generally, each reaction needs an enzyme. The metabolic pathway collects a series of chemical reactions at the system level. As compounds and enzymes are two important components in each metabolic pathway, identification of metabolic pathways that a given compound or enzyme can participate is the first important step for understanding the mechanism of metabolic pathways. Objective: The purpose of this study was to build efficient computational methods to predict the metabolic pathways of compounds and enzymes. Methods: Novel multi-label classifiers were proposed to identify metabolic pathway types, reported in KEGG, of compounds and enzymes. Three heterogeneous networks defining compounds and enzymes as nodes were constructed. To extract more informative features of compounds and enzymes, we generalized the powerful network embedding algorithm, Mashup, to its heterogeneous network version, named MashupH. RAndom k-labELsets (RAKEL) was employed to build the classifiers and support vector machine or random forest was selected as the base classification algorithm. Results: The 10-fold cross-validation results indicated the good performance of the proposed classifiers and such performance was superior to the previous classifier that adopted features yielded by Mashup. Furthermore, some key parameters of MashupH that might contribute to or influence the classifiers were analyzed. Conclusion: The features yielded by MashupH were more informative than those produced by Mashup on heterogeneous networks. This was the main reason the new classifiers were superior to those using features yielded by Mashup.

Fast Solution-Phase and Liquid-Phase Peptide Syntheses (SolPSS and LPPS) Mediated by Biomimetic Cyclic Propylphosphonic Anhydride (T3P®).

The growing applications of peptide-based therapeutics require the development of efficient protocols from the perspective of an industrial scale-up. T3P® (cyclic propylphosphonic anhydride) promotes amidation in the solution-phase through a biomimetic approach, similar to the activation of carboxylic moiety catalyzed by ATP-grasp enzymes in metabolic pathways. The T3P® induced coupling reaction was applied in this study to the solution-phase peptide synthesis (SolPPS). Peptide bond formation occurred in a few minutes with high efficiency and no epimerization, generating water-soluble by-products, both using N-Boc or N-Fmoc amino acids. The optimized protocol, which was successfully applied to the iterative synthesis of a pentapeptide, also allowed for a decrease in the solvent volume, thus improving process sustainability. The protocol was finally extended to the liquid-phase peptide synthesis (LPPS), where the isolation of the peptide was performed using precipitation, thus also showing the suitability of this coupling reagent to this emerging technique.

Characterization of the glucuronidating pathway of pectolinarigenin, the major active constituent of the Chinese medicine Daji, in humans and its influence on biological activities

Ethnopharmacological relevanceThe Chinese medicine Daji (the aerial part of Cirsium japonicum DC.) and its charred product (Cirsii Japonici Herba Carbonisata) have been widely used as hemostatic agents or diuretic agents to prepare a variety of Chinese herbal formula. Pectolinarigenin (PEC), one of the most abundant constituents in both Daji and its charred product, has been considered as the key effective substance responsible for the major pharmacological activities of Daji, including hemostasis, hepatoprotective, anti-tumor and anti-osteoporosis effects. However, the major metabolic pathways of PEC in humans and the influence of PEC metabolism on its biological activities are poorly understood. Aim of the studyTo characterize the main metabolic pathway(s) and key enzymes of PEC in human biological systems, as well as to reveal the influence of PEC metabolism on its biological activities. Materials and methodsThe metabolic stability assays of PEC were investigated in human liver microsomes (HLM). The O-glucuronide of PEC was biosynthesized and characterized by nuclear magnetic resonance (NMR) spectroscopy. The key enzymes responsible for O-glucuronidation of PEC in humans were assigned by performing UGT reaction phenotyping, chemical inhibition and enzymatic kinetic assays. The agonist effects of PEC and its O-glucuronide on nuclear factor erythroid2-related factor 2 (Nrf2), Peroxisome proliferator activated receptors (PPARα and PPARβ) were tested at the cellular level. ResultsPEC could be readily metabolized to form a mono-O-glucuronide in both human liver microsome (HLM) and human intestinal microsome (HIM). The mono-O-glucuronide was bio-synthesized by mouse liver S9 and its structure was fully characterized as PEC-7-O-β-D-glucuronide (PEC-O-7-G). UGT1A1, UGT1A3 and UGT1A9 are key enzymes responsible for PEC-7-O-glucuronidation in HLM, while UGT1A1, UGT1A9 and 1A10 may play key roles in this reaction in HIM. Biological tests revealed that PEC displayed strong agonist effects on Nrf2, PPARα and PPARβ, whereas PEC-7-O-glucuronide showed relatively weak Nrf2 agonist effect and very weak PPAR agonist effects, indicating that PEC-7-O-glucuronidation strongly weaken its agonist effects on Nrf2 and PPAR. ConclusionsOur results demonstrate that 7-O-glucuronidation is the major metabolic pathway of PEC in human tissues, while UGT1A1, 1A3 and 1A9 are key contributing enzymes responsible for PEC-7-O-glucuronidation in human liver. It is also found that PEC 7-O-glucuronidation significantly weakens the Nrf2 and PPAR agonist effects. All these findings are very helpful for the pharmacologists to deep understand the metabolic rates of PEC in humans.

Comparative proteomics unveils the bacteriostatic mechanisms of Ga(III) on the regulation of metabolic pathways in Pseudomonas aeruginosa

Gallium has a long history as a chemotherapeutic agent. The mechanisms of action of Ga(III)-based anti-infectives are different from conventional antibiotics, which primarily result from the chemical similarities of Ga(III) with Fe(III) and substitution of gallium into iron-dependent biological pathways. However, more aspects of the molecular mechanisms of Ga(III) against human pathogens, especially the effects on bacterial metabolic processes, remain to be understood. Herein, by using conventional quantitative proteomics, we identified the protein changes of Pseudomonas aeruginosa (P. aeruginosa) in response to Ga(NO3)3 treatment. We show that Ga(III) exhibits bacteriostatic mode of action against P. aeruginosa through affecting the expressions of a number of key enzymes in the main metabolic pathways, including glycolysis, TCA cycle, amino acid metabolism, and protein and nucleic acid biosynthesis. In addition, decreased expressions of proteins associated with pathogenesis and virulence of P. aeruginosa were also identified. Moreover, the correlations between protein expressions and metabolome changes in P. aeruginosa upon Ga(III) treatment were identified and discussed. Our findings thus expand the understanding on the antimicrobial mechanisms of Ga(III) that shed light on enhanced therapeutic strategies. Biological significanceMounting evidence suggest that the efficacy and resistance of clinical antibiotics are closely related to the metabolic homeostasis in bacterial pathogens. Ga(III)-based compounds have been repurposed as antibacterial therapeutic candidates against antibiotics resistant pathogens, and represent a safe and promising treatment for clinical human infections, while more thorough understandings of how bacteria respond to Ga(III) treatment are needed. In the present study, we provide evidences at the proteome level that indicate Ga(III)-induced metabolic perturbations in P. aeruginosa. We identified and discussed the interference of Ga(III) on the expressions and activities of enzymes in the main metabolic pathways in P. aeruginosa. In view of our previous report that the antimicrobial efficacy of Ga(III) could be modulated according to Ga(III)-induced metabolome changes in P. aeruginosa, our current analyses may provide theoretical basis at the proteome level for the development of efficient gallium-based therapies by exploiting bacterial metabolic mechanisms.

Effects of lysed sludge reflux point on ultrasound lysis-cryptic growth in anaerobic/aerobic (A/O) wastewater treatment: Sludge reduction, microbial community, and metabolism

Ultrasonication allows sludge reduction to be performed in situ during wastewater treatment, and the reflux point of the lysed sludge affects this performance. This study investigated the effects of reflux point (anaerobic stage, carbon/nitrogen (C/N) lowest stage, and aerobic stage) on sludge lysis-cryptic growth in an anaerobic/aerobic reactor and variations in the sludge and microbial community. The best reflux point occurred at the lowest C/N ratio stage, and a 50.96% reduction in excess sludge was achieved. The reflux of the lysed sludge to the aerobic stage reduced nitrogen and phosphorus removal. The reflux of the lysed sludge decreased the average sludge size, reaching 29.2 μm when reflux to the aerobic stage. Scanning electron microscopy showed that the sludge surface was unaffected by the reflux point. The Fourier-transform infrared spectrometry and X-ray photoelectron spectroscopy results showed that the most prominent variation in the intensity of the sludge functional groups occurred when the reflux was at the lowest C/N stage. The amount of extracellular polymeric substances decreased the most during reflux to the anaerobic stage. The sludge microbial communities varied with the reflux point, and the dominant phyla during reflux to the anaerobic, lowest C/N, and aerobic stages were Bacteroidetes, Firmicutes, and Bacteroidetes, respectively. Furthermore, the reflux point did not alter the metabolic pathway of sludge microorganisms but increased the number of enzymes in metabolic pathways.

Integrating metabolomics, 16S rRNA sequencing, network pharmacology, and metorigin to explore the mechanism of Cinnamomi Cortex in treating chronic atrophic gastritis rats

BackgroundCinnamomi cortex called as Rougui (RG) in Chinese was a widely used food-medicine homology. RG has the potential to treat chronic atrophic gastritis (CAG), a disease with widespread impact in the Chinese population. PurposeThis study aimed to explore its mechanism against CAG based on amalgamated strategies. MethodsNetwork pharmacology was used to predict the potential effective components and the core targets of RG against CAG based on the comprehensive chemical characterization using UHPLC-Q/TOF MS (ultra high performance liquid chromatogramphy-quadrupole/time-of-flight mass spectrometry). The CAG animals model were further used to validate its pharmacodynamics, of which gut microbiota of caecal contents were analyzed by integrating metabolomics, 16S rRNA sequencing, Metorigin metabolite traceability analysis and molecular docking to explore its action mechanism. ResultsNetwork pharmacology firstly predicted the efficacy of RG was attributed to four effective components and seven targets. Metabolomics of caecal contents in CAG rats revealed primary bile acid biosynthesis was its targeted metabolic pathway associated with the metabolism of gut microbiota coupled with Metorigin traceability analysis. 16S rRNA sequencing showed that RG treated CAG by regulating the imbalance of gut microbiota. Molecular docking further confirmed that the effective components of RG could intervene with potential targets, metorigin analysis pathway, and key enzymes of gut microbiota metabolic pathways. ConclusionOur results proved that RG exerted favorable effect on CAG. The four active ingredients (quercetin, kaempferol, oleic acid, and (-)-epicatechin) of RG were the key to exert drug effect, which could targeted the core target of CAG, primary bile acid biosynthesis and intestinal flora metabolic pathways.