FBPase Activity Research Articles

Herbacetin Inhibits Human Fructose 1,6-Bisphosphatase Among a Panel of Chromone Derivatives and Pyrazoles, Demonstrating Positive Effects on Insulin-Resistant HepG2 Cells.

In patients with type 2 diabetes mellitus (DM), excessive gluconeogenesis is considered a major contributor to hyperglycemia. Therefore, targeting fructose 1,6-bisphosphatase (FBPase), a key regulatory enzyme involved in gluconeogenesis, has gained interest as a potential therapeutic target for managing DM. In this study, a library of 42 structurally-related chromone derivatives (including flavonoids, 2-styrylchromones, and 2-(4-arylbuta-1,3-dien-1-yl)chromones, named as 2-styrylchromone-related derivatives), as well as 4- and 5-styrylpyrazoles, were tested against human FBPase using a noncellular microanalysis screening system. Herbacetin, 3,4',5,7,8-pentahydroxyflavone, inhibited FBPase activity with an IC50 value of 6.4 ± 0.7 μM. The effects of herbacetin were also explored using an insulin-resistant human hepatocellular carcinoma cell line (HepG2 cells). The results showed that herbacetin significantly decrease insulin resistance by promoting the phosphorylation of protein kinase B (Akt), and exhibited a capacity to ameliorate inflammation, evidenced by the modulation of the inhibitor of κB alpha (IκBα). This study provides important considerations for the design of novel FBPase inhibitors. Furthermore, it indicates a preliminary potential of herbacetin's dual action in improving insulin resistance and decreasing inflammation, suggesting the need for further investigation of this compound for addressing the complexities of type 2 DM management.

Mitochondria targeted esculetin administration improves insulin resistance and hyperglycemia-induced atherosclerosis in db/db mice.

The development and progression of hyperglycemia (HG) and HG-associated atherosclerosis are exacerbated by mitochondrial dysfunction due to dysregulated mitochondria-derived ROS generation. We recently synthesized a novel mitochondria-targeted esculetin (Mito-Esc) and tested its dose-response therapeutic efficacy in mitigating HG-induced atherosclerosis in db/db mice. In comparison to simvastatin and pioglitazone, Mito-Esc administration resulted in a considerable reduction in body weights and improved glucose homeostasis, possibly by reducing hepatic gluconeogenesis, as indicated by a reduction in glycogen content, non-esterified free fatty acids (NEFA) levels, and fructose 1,6-bisphosphatase (FBPase) activity. Interestingly, Mito-Esc treatment, by regulating phospho-IRS and phospho-AKT levels, greatly improved palmitate-induced insulin resistance, resulting in enhanced glucose uptake in adipocytes and HepG2 cells. Also, and importantly, Mito-Esc administration prevented HG-induced atheromatous plaque formation and lipid accumulation in the descending aorta. In addition, Mito-Esc administration inhibited the HG-mediated increase in VACM, ICAM, and MAC3 levels in the aortic tissue, as well as reduced the serum pro-inflammatory cytokines and markers of senescence. In line with this, Mito-Esc significantly inhibited monocyte adherence to human aortic endothelial cells (HAECs) treated with high glucose and reduced high glucose-induced premature senescence in HAECs by activating the AMPK-SIRT1 pathway. In contrast, Mito-Esc failed to regulate high glucose-induced endothelial cell senescence under AMPK/SIRT1-depleted conditions. Together, the therapeutic efficacy of Mito-Esc in the mitigation of hyperglycemia-induced insulin resistance and the associated atherosclerosis is in part mediated by potentiating the AMPK-SIRT1 axis. KEY MESSAGES: Mito-Esc administration significantly mitigates diabetes-induced atherosclerosis. Mito-Esc improves hyperglycemia (HG)-associated insulin resistance. Mito-Esc inhibits HG-induced vascular senescence and inflammation in the aorta. Mito-Esc-mediated activation of the AMPK-SIRT1 axis regulates HG-induced endothelial cell senescence.

Excess homocysteine inhibits pancreatic β-cell secretory function by repressing Zbtb20 expression

Homocysteine (Hcy) is a sulfur-containing amino acid. An elevated level of Hcy is a risk factor for diabetes development. However, the mechanism of its effect on pancreatic β-cell function is unclear. In this study, we constructed a hyperhomocysteinemia (HHcy) mouse model by feeding mice a high methionine diet (HMD). The mice suffered impaired glucose tolerance and reduced insulin secretion. Furthermore, at the cellular level, INS1 cells exhibited impaired insulin secretory function after the Hcy intervention. Transcriptomics revealed that Zbtb20 expression was downregulated and the downstream gene Fbp1 was upregulated in HHcy-induced mice compared with mice fed with normal diet. Insulin secretion could be restored by Zbtb20 overexpression or fructose 1,6-bisphosphatase (FBPase) activity inhibition in INS1 cells. In conclusion, our study suggested that Hcy inhibited the insulin secretory function of pancreatic β-cells by suppressing Zbtb20 expression, leading to the development of diabetes. Zbtb20 may be a key target in the development of diabetes associated with elevated Hcy levels.

Effects of Xanthomonas campestris pv. campestris on the photosynthesis of cabbage in the early stage of infection

Cabbage is one of the most important cruciferous vegetables. Black rot, caused by Xanthomonas campestris pv. campestris (Xcc) seriously affects cabbage production; therefore, it is important to study the resistance mechanism to black rot in cabbage. In this study, cabbage photosynthetic physiology changes in the early stages of Xcc infection were analyzed using physiological index determination and transcriptome analysis. The results showed that the Xcc infection can lead the chlorophyll degradation and decreased the net photosynthetic rate (Pn), and the decrease in Pn was mainly due to non-stomatal factors. In addition, the maximum photosynthetic efficiency (Fv/Fm), actual photosynthetic efficiency of photosystem II (PSII) gradually decreased, and the OJIP curve tended to be flat, indicating that Xcc infection reduced the activity of the PSII reaction center and affected the chlorophyll fluorescence intensity. Moreover, the primary photochemical reaction of cabbage leaves was damaged, and electron transfer on the PSII receptor side was inhibited after Xcc infection. NADP-GAPDH and FBPase activities in the Calvin cycle decreased after Xcc invasion. Using transcriptome analysis, 173, 77, 63, and 53 differentially expressed genes (DEGs) were enriched in "carbon metabolism", "photosynthesis", "carbon fixation in photosynthetic organisms", and "metabolism of porphyrins and chlorophyll" metabolic processes, respectively. These results suggest that Xcc infection affects cabbage photosynthesis via multiple pathways. Our results provide a theoretical basis for further research on black rot-resistant breeding and cabbage resistance mechanisms.

Virtual screening, molecular docking, molecular dynamics, and MM-GBSA approaches identify prospective fructose-1,6-bisphosphatase inhibitors from pineapple for diabetes management

Diabetes affects millions globally and poses treatment challenges. Targeting the enzyme fructose-1,6-bisphosphatase (FBPase) in gluconeogenesis and exploring plant-based therapies offer potential solutions for improving diabetes management while supporting sustainability and medicinal advancements. Utilizing pineapple (Ananas comosus L. Merr.) waste as a source of drug precursors could be valuable for health and environmental care due to its medicinal benefits and abundant yearly biomass production. Therefore, this study conducted a virtual screening to identify potential natural compounds from pineapple that could inhibit FBPase activity. A total of 112 compounds were screened for drug-likeness and ADMET properties, and molecular docking simulations were performed on 20 selected compounds using blind docking. The lead compound, butane-2,3-diyl diacetate, was subjected to 100 ns MD simulations, revealing a binding energy of −5.4 kcal/mol comparable to metformin (−5.6 kcal/mol). The MD simulation also confirmed stable complexes with crucial hydrogen bonds. Glu20, Ala24, Thr27, Gly28, Glu29, Leu30, Val160, Met177, Asp178, and Cys179 were identified as key amino acids that stabilized the human liver FBPase-butane-2,3-diyl diacetate complex, while Tyr215 and Asp218 played a crucial role in the human liver FBPase-Metformin complex. Our study indicates that the lead compound has high intestinal solubility. Therefore, it would show rapid bloodstream distribution and effective action on the target protein, making butane-2,3-diyl diacetate a potential antidiabetic drug candidate. However, further investigations in vitro, preclinical, and clinical trials are required to thoroughly assess its efficacy and safety. Communicated by Ramaswamy H. Sarma

An efficient triose phosphate synthesis and distribution in wheat provides tolerance to higher field temperatures.

High temperatures in the field hinder bread wheat high-yield production, mainly because of the adverse effects of heat over photosynthesis. The Yaqui Valley, the main wheat producer region in Mexico, is a zone prone to have temperatures over 30°C. The aim of this work was to test the flag leaf photosynthetic performance in 10 bread wheat genotypes grown under high temperatures in the field. The study took place during two seasons (2019-2020 and 2020-2021). In each season, control seeds were sown in December, while heat-stressed were sown in late January to subject wheat to heat stress (HS) during the grain-filling stage. HS reduced Grain yield from 20 to 58% in the first season. HS did not reduce chlorophyll content and light-dependent reactions were unaffected in any of the tested genotypes. Rubisco, chloroplast fructose 1,6-biphosphatase (FBPase), and sucrose phosphate synthase (SPS) activities were measured spectrophotometrically. Rubisco activity did not decrease under HS in any of the genotypes. FBPase activity was reduced by HS indicating that triose phosphate flux to starch synthesis was reduced, while SPS was not affected, and thus, sucrose synthesis was maintained. HS reduced aerial biomass in the 10 chosen genotypes. Genotypes SOKWB.1, SOKWB.3, and BORLAUG100 maintained their yield under HS, pointing to a potential success in their introduction in this region for breeding heat-tolerant bread wheat.

Wheat Transformation with ScTPS1-TPS2 Bifunctional Enzyme for Trehalose Biosynthesis Protects Photosynthesis during Drought Stress

Wheat cultivation makes an important contribution to human nutrition. Trehalose synthesis plays a role in the tolerance to drought stress. A bifunctional TPS-TPP enzyme gene from yeast was used to obtain transgenic wheat plants to increase trehalose synthesis. Mature wheat embryos were transformed using pGreen rd29A::TPS1-TPS2 or pGreen 35S::TPS1-TPS2 constructs. The transgene presence in mature leaves of T3 plants was confirmed by sequencing a PCR fragment of the inserted transgene. Transgenic and NT plants were submitted to drought stress for eight days. Transformed wheat lines retained a higher relative water content than NT plants during drought stress, and the Rubisco activity was unaffected. Plants transformed with the 35S construct showed a lower photosynthetic rate and lower fructose 1–6-bisphosphatase (FBPase) activity during drought, suggesting that constitutive trehalose and sucrose synthesis caused a reduced ribulose 1,5-bisphosphate (RuBP) regeneration. Lines transformed with the rd29A promoter showed a higher photosynthetic rate after eight days of drought, as the RuBP regeneration was unaffected. Transgenic wheat plants had higher biomass and grain weight than NT plants after drought. These results suggest that trehalose synthesis improves photosynthesis during stress and induces changes in the activity of some Calvin-cycle enzymes, reflected in plant metabolism and growth.

Effects of CO2 Enrichment on Carbon Assimilation, Yield and Quality of Oriental Melon Cultivated in a Solar Greenhouse

Since CO2 is the fundamental substrate for photosynthesis, fluctuating concentrations have a direct effect on plant growth and metabolism. Accordingly, CO2 enrichment within a certain range was found to improve photosynthesis, yields and the quality of plants. In order to further understand the underlying impact of CO2 enrichment, this study employed an open-top chamber growth box model with the following two treatments: control treatment (CO2 concentration: 380 ± 30 μL/L) and CO2 enrichment (1200 ± 50 μL/L). The effects on leaf carbon assimilation, fruit yield and quality were subsequently determined. The net photosynthetic rate, intercellular CO2 concentration, dry matter accumulation and soluble sugar content in the oriental melon leaves increased significantly on day 5 of CO2 enrichment. Moreover, a significant increase in the activity of carbon assimilation-related enzymes Rubisco, RCA, FBPase and CA was also observed, with the upregulation of CmRubisco, CmRCA, CmFBPase and CmCA gene expression from day 15 of CO2 enrichment. Thus, the yield per plant and content of soluble sugars and soluble solids in the fruit also increased significantly. These findings suggest that CO2 enrichment has positive effects on oriental melon growth, increasing photosynthesis and the activity of photosynthetic carbon-assimilation-related enzymes and associated gene expression, thereby improving fruit yields and quality. These results provide a foundation for the CO2 enrichment of oriental melon cultivated in solar greenhouses in autumn/winter and winter/spring.

Effects of Dietary Carbohydrate Sources on Growth Performance, Glucose Metabolism, and Digestive Enzyme Activity of Litopenaeus vannamei

This study evaluated the effects of different sources of dietary carbohydrates on the growth, antioxidant capacity, and digestive enzyme activity, and carbohydrate metabolic enzyme activity of Litopenaeus vannamei were evaluated. The experimental diets, including glucose, sucrose, potato starch, corn starch, and wheat starch, were fed to L. vannamei for 35 days. We found that shrimp fed on corn starch and wheat starch grew better, whereas shrimp fed on wheat starch showed the highest digestive enzyme activity. In addition, the starch group showed higher total superoxide dismutase (T-SOD) activity and lower malondialdehyde (MDA) content. The glucose, sucrose, starch, and corn starch fed shrimp showed significantly higher levels of hepatopancreatic hexokinase (HK), whereas those fed on glucose and potato starch also showed significantly higher levels of hepatopancreatic phosphofructokinase (PFK) activity. The activities of phosphoenolpyruvate carboxykinase (PEPCK) and glucose 6-phosphatase (G6Pase) were higher in shrimp fed on glucose, whereas higher fructose 1,6-bisphosphatase (FBPase) activity was observed in shrimp fed on wheat starch, glucose, and sucrose. The insulin content of shrimp in the glucose group and potato starch group was higher, whereas that of the glucose group was higher than that of the potato starch group. In brief, the addition of wheat starch to the diet plays a positive role in the growth, glycolysis, gluconeogenesis, and antioxidant activity of shrimp, and is a promising choice for juvenile shrimp culture.

Subcellular compartmentalization of aluminum reduced its hazardous impact on rye photosynthesis

Aluminum (Al) toxicity limits crops growth and production in acidic soils. Compared to roots, less is known about the toxic effects of Al in leaves. Al subcellular compartmentalization is also largely unknown. Using rye (Secale cereale L.) Beira (more tolerant) and RioDeva (more sensitive to Al) genotypes, we evaluated the patterns of Al accumulation in leaf cell organelles and the photosynthetic and metabolic changes to cope with Al toxicity. The tolerant genotype accumulated less Al in all organelles, except the vacuoles. This suggests that Al compartmentalization plays a role in Al tolerance of Beira genotype. PSII efficiency, stomatal conductance, pigment biosynthesis, and photosynthesis metabolism were less affected in the tolerant genotype. In the Calvin cycle, carboxylation was compromised by Al exposure in the tolerant genotype. Other Calvin cycle-related enzymes, phoshoglycerate kinase (PGK), glyceraldehyde 3-phosphate dehydrogenase (GAPDH), triose-phosphate isomerase (TPI), and fructose 1,6-bisphosphatase (FBPase) activities decreased in the sensitive line after 48 h of Al exposure. Consequentially, carbohydrate and organic acid metabolism were affected in a genotype-specific manner, where sugar levels increased only in the tolerant genotype. In conclusion, Al transport to the leaf and compartmentalization in the vacuoles tolerant genotype's leaf cells provide complementary mechanisms of Al tolerance, protecting the photosynthetic apparatus and thereby sustaining growth.

Biocontrol potential of Trichoderma harzianum against Botrytis cinerea in tomato plants

Trichoderma harzianum (TM) is a soil-borne beneficial fungus that positively affects plant growth and defense. TM shows antagonism against multiple plant pathogens; however, the TM as a biocontrol agent against Botrytis cinerea (BC) and underlying mechanisms remain unclear. Herein we studied the effects of TM on BC by dual-culture assay and greenhouse bioassay in tomato plants. In vitro experiments showed that TM rapidly occupied most of the nutrient space and eventually covered BC on potato dextrose agar (PDA) medium. The TM inhibition rate against BC was 62.05% on the sixth day of culture. The inhibitory effect of TM spore suspension on gray mold inoculated with spore suspension and solid inoculants was 33.66% and 69.44%, respectively in tomato plants. Notably, TM decreased the leaf concentrations of BC-induced hydrogen peroxide and malondialdehyde but improved the redox state. TM inoculation caused biphasic peaks in the expression of secondary metabolism and defense-related genes and increased the concentrations of flavonoids, phenols and lignin in tomato leaves. Moreover, TM alleviated stomatal closure and enhanced the net photosynthesis rate and the activity of RuBisCO and FBPase compared with only BC inoculation. Our results suggest that TM spore suspension improves tomato resistance to gray mold disease by stimulating cellular redox, secondary metabolites and stomatal movement.

New Insights to Regulation of Fructose-1,6-bisphosphatase during Anoxia in Red-Eared Slider, Trachemys scripta elegans.

The red-eared slider (Trachemys scripta elegans) undergoes numerous changes to its physiological and metabolic processes to survive without oxygen. During anoxic conditions, its metabolic rate drops drastically to minimize energy requirements. The alterations in the central metabolic pathways are often accomplished by the regulation of key enzymes. The regulation of one such enzyme, fructose-1,6-bisphosphatase (FBPase; EC 3.1.3.11), was characterized in the present study during anoxia in liver. FBPase is a crucial enzyme of gluconeogenesis. The FBPase was purified from liver tissue in both control and anoxic conditions and subsequently assayed to determine the kinetic parameters of the enzyme. The study revealed the relative degree of post-translational modifications in the FBPase from control and anoxic turtles. Further, this study demonstrated a significant decrease in the maximal activity in anoxic FBPase and decreased sensitivity to its substrate Fructose-1,6-bisphosphate (FBP) when compared to the control. Immunoblotting demonstrated increased threonine phosphorylation (~1.4-fold) in the anoxic FBPase. Taken together, these results suggest that the phosphorylation of liver FBPase is an important step in suppressing FBPase activity, ultimately leading to the inhibition of gluconeogenesis in the liver of the red-eared slider during anaerobic conditions.

The physiological and proteomic characteristics of oilseed rape stem affect seed yield and lodging resistance under different planting densities and row spacing

AbstractLodging is an important factor that restricts the yield of rapeseed and the mechanical harvesting. To explore agricultural practise and mechanisms that can synergistically improve the rapeseed yield and lodging resistance, a conventional plant type HZ 62 was selected in 2016–2017, and HZ 62 and an erect‐leaf type 1,301 were grown in 2017–2018. The planting density was set as main plots at 15 × 104 plants ha1 (D1), 30 × 104 plants ha‐1 (D2) and 45 × 104 plants ha−1 (D3), with the row spacing as sub‐plots of 15 cm (R15), 25 cm (R25) and 35 cm (R35). The results showed that with increased density and narrowed row spacing, the radiation utilisation efficiency (RUE) of the population and seed yield were greatly increased. Compared with that at D1R25, the yield of HZ 62 at D3R15 was increased by 19.00% and 22.53%, respectively, in 2016–2017 and 2017–2018, whereas the yield of 1,301 rose by 31.98%. The influence of the RUE on the seed yield reached the peak at flowering stage. With increased planting density and narrowed row spacing, the lodging index of HZ 62 declined. The lodging index of 1,301 was increased with higher density and declined with narrowed row spacing. Compared with D1R25, the averaged lodging index across different parts of HZ 62 at D3R15 declined by 15.03% and 10.80%, respectively, in 2016–2017 and 2017–2018, and that of 1,301 was increased by 25.47%. Compared with D1R25, the stem proteomics during the flowering stage showed the expression of photosynthesis‐related proteins (A0A078FHG9, A0A078JZM9 and A0A078JQI2) up‐regulated, that consistent with Rubisco, SPS and FBPase activities; proteins related to phenylpropanes metabolism, A0A078IWR4 of HZ 62 and A0A078JWZ9 of 1,301 increased that consistent with POD activity, which resulted in more sucrose, starch and lignin in the stem at D3R15. Moreover, the protein related to cellulose synthesis (A0A078I4D9), the activity of β‐1,3‐glucanase and content of cellulose in the stem were up‐regulated in HZ 62 but down‐regulated in 1,301, contrary to the variation in the stem lodging index. The cellulose synthesis differed between the varieties in different treatment combinations of planting density and row spacing. This was the key process by which planting density and row spacing influence the lodging of rapeseed varieties with varied plant architecture.

Melatonin-mediated photosynthetic performance of tomato seedlings under high-temperature stress

Photosynthesis is a fundamental biosynthetic process in plants that can enhance carbon absorption and increase crop productivity. Heat stress severely inhibits photosynthetic efficiency. Melatonin is a bio-stimulator capable of regulating diverse abiotic stress tolerances. However, the underlying mechanisms of melatonin-mediated photosynthesis in plants exposed to heat stress largely remain elucidated. Our results revealed that melatonin treatment (100 μM) in tomato seedlings increased the endogenous melatonin levels and photosynthetic pigment content along with upregulated of their biosynthesis gene expression under high-temperature stress (42 °C for 24 h), whereas heat stress significantly decreased the values of gas exchange parameters. Under heat stress, melatonin boosted CO2 assimilation, i.e., Vc,max (maximum rate of ribulose-1,5-bisphosphate carboxylase, Rubisco), and Jmax (electron transport of Rubisco generation) and also enhanced the Rubisco and FBPase activities, which resulted in upregulated photosynthetic related gene expression. In addition, heat stress greatly reduced the photochemical chemistry of photosystem II (PSII) and photosystem I (PSI), particularly the maximum quantum efficiency of PSII (Fv/Fm) and PSI (Pm). Conversely, melatonin supplementation increased the chlorophyll a fluorescence parameters led to amplifying the electron transport efficiency. Moreover, heat stress decreased the actual PSII efficiency (ΦPSII), electron transport rate (ETR) and photochemical quenching coefficient (qP), while increasing nonphotochemical quenching (NPQ); however, melatonin reversed these values, which helps to fostering the dissipation of excess excitation energy. Taken together, our results provide a concrete insight into the efficacy of melatonin-mediated photosynthesis performance in a high-temperature regime.

Regulation of Fructose 1,6-Bisphosphatase in Procyclic Form Trypanosoma brucei.

Glycolysis is well described in Trypanosoma brucei, while the importance of gluconeogenesis and one of the key enzymes in that pathway, fructose 1,6-bisphosphatase, is less understood. Using a sensitive and specific assay for FBPase, we demonstrate that FBPase activity in insect stage, procyclic form (PF), parasite changes with parasite cell line, extracellular glucose levels, and cell density. FBPase activity in log phase PF 2913 cells was highest in high glucose conditions, where gluconeogenesis is expected to be inactive, and was undetectable in low glucose, where gluconeogenesis is predicted to be active. This unexpected relationship between FBPase activity and extracellular glucose levels suggests that FBPase may not be exclusively involved in gluconeogenesis and may play an additional role in parasite metabolism. In stationary phase cells, the relationship between FBPase activity and extracellular glucose levels was reversed. Furthermore, we found that monomorphic PF 2913 cells had significantly higher FBPase levels than pleomorphic PF AnTat1.1 cells where the activity was undetectable except when cells were grown in standard SDM79 media, which is glucose-rich and commonly used to grow PF trypanosomes in vitro. Finally, we observed several conditions where FBPase activity changed while protein levels did not, suggesting that the enzyme may be regulated via post-translational modifications.

Research on Drug Molecular Computing Based on Support Vector Machine and Neural Network

With the continuous development of science, a large number of drugs, molecules and genes have been stored in various databases. The relationship between drug like molecules and molecular properties can be identified by computer science methods according to the diversity of compound library, drug molecular characteristics and the differences between different molecules, so as to achieve the purpose of drug screening. In this paper, 300 FBPase inhibitors were selected from previous literatures to convert lC50 into plC50. The whole data set is divided into 236 molecules of training set and 64 molecules of test set. Two machine learning algorithms, namely Neural Network and Support Vector Machine, are selected to construct the prediction model with the training set as the learning object. And the reliability of the model is verified by the test set, and it is applied to the prediction of FBPase inhibitors. The prediction results show that the model is stable and reliable. In addition, the randomization test shows that the current model is not caused by chance correlation. The explanation of the selected molecular descriptors proves that the polarity of the molecule plays an important role in the inhibitory activity of FBPase. The model can provide some useful guidance for drug researchers and can screen ideal drug molecules before drug development.

Discovery of N-Arylsulfonyl-Indole-2-Carboxamide Derivatives as Potent, Selective, and Orally Bioavailable Fructose-1,6-Bisphosphatase Inhibitors-Design, Synthesis, In Vivo Glucose Lowering Effects, and X-ray Crystal Complex Analysis.

Liver fructose-1,6-bisphosphatase (FBPase) is a key enzyme in the gluconeogenesis pathway. Inhibiting FBPase activity represents a potential treatment for type 2 diabetes mellitus. A series of novel N-arylsulfonyl-4-arylamino-indole-2-carboxamide derivatives have been disclosed as FBPase inhibitors. Through extensive structure-activity relationship investigations, a promising candidate molecule Cpd118 [sodium (7-chloro-4-((3-methoxyphenyl)amino)-1-methyl-1H-indole-2-carbonyl] [(4-methoxyphenyl)sulfonyl)amide] has been identified with high inhibitory activity against human liver FBPase (IC50, 0.029 ± 0.006 μM) and high selectivity relative to the other six AMP-binding enzymes. Importantly, Cpd118 produced significant glucose-lowering effects on both type 2 diabetic KKAy mice and ZDF rats as demonstrated by substantial reductions in the fasting and postprandial blood glucose levels, as well as the HbA1c level. Furthermore, Cpd118 elicited a favorable pharmacokinetic profile with an oral bioavailability of 99.1%. Moreover, the X-ray crystal structure of the Cpd118-FBPase complex was resolved, which revealed a unique binding mode and provided a structural basis for its high potency and selectivity.

Biological evaluation and SAR analysis of novel covalent inhibitors against fructose-1,6-bisphosphatase

Fructose-1,6-bisphosphatase (FBPase) is an attractive target for affecting the GNG pathway. In our previous study, the C128 site of FBPase has been identified as a new allosteric site, where several nitrovinyl compounds can bind to inhibit FBPase activity. Herein, a series of nitrostyrene derivatives were further synthesized, and their inhibitory activities against FBPase were investigated in vitro. Most of the prepared nitrostyrene compounds exhibit potent FBPase inhibition (IC50 < 10 μM). Specifically, when the substituents of F, Cl, OCH3, CF3, OH, COOH, or 2-nitrovinyl were installed at the R2 (meta-) position of the benzene ring, the FBPase inhibitory activities of the resulting compounds increased 4.5–55 folds compared to those compounds with the same groups at the R1 (para-) position. In addition, the preferred substituents at the R3 position were Cl or Br, thus compound HS36 exhibited the most potent inhibitory activity (IC50 = 0.15 μM). The molecular docking and site-directed mutation suggest that C128 and N125 are essential for the binding of HS36 and FBPase, which is consistent with the C128-N125-S123 allosteric inhibition mechanism. The reaction enthalpy calculations show that the order of the reactions of compounds with thiol groups at the R3 position is Cl > H > CH3. CoMSIA analysis is consistent with our proposed binding mode. The effect of compounds HS12 and HS36 on glucose production in primary mouse hepatocytes were further evaluated, showing that the inhibition was 71% and 41% at 100 μM, respectively.

Identification of the New Covalent Allosteric Binding Site of Fructose-1,6-bisphosphatase with Disulfiram Derivatives toward Glucose Reduction.

Fructose 1,6-bisphosphatase (FBPase) has attracted substantial interest as a target associated with cancer and type 2 diabetes. Herein, we found that disulfiram and its derivatives can potently inhibit FBPase by covalently binding to a new C128 allosteric site distinct from the original C128 site in APO FBPase. Further identification of the allosteric inhibition mechanism reveals that the covalent binding of a fragment of 214 will result in the movement of C128 and the dissociation of helix H4 (123-128), which in turn allows S123 to more easily form new hydrogen bonds with K71 and D74 in helix H3 (69-72), thereby inhibiting FBPase activity. Notably, both disulfiram and 212 might moderately reduce blood glucose output in vivo. Therefore, our current findings not only identify a new covalent allosteric site of FBPase but also establish a structural foundation and provide a promising way for the design of covalent allosteric drugs for glucose reduction.

Brassinosterioids Promote Photosynthetic Parameters of Pigeon Pea Plants under Water Deficit Conditions

This study was aimed to find the effects of 28-epibrassinolide (28-EBL) and HBL on pigeon pea seedlings subjected to drought stress, either alone and supplemented with 28-EBL and HBL treatments. Supplementation of EBL alone also exhibited the significant improvement on chlorophyll content (44.3% of Chl a and 54.3 % of Chl b) than EBL under drought stress treatments compared to the control plants. Control plants receiving the EBL alone treatment showed the significant effect (by 36%) than the EBL under stress treatment (19.78%) compared to the control for carotenoid levels. supplementation of EBL exhibited the significant improvement of PN (36%; 0.0411, p≤0.05) over their individuals in comparison to control. application of EBL under unstressed was more effective (45%; 0.0341, p≤0.05) over stressed control than their individual applications in enhancing gS in control plants. Plants treated with HBL alone showed a marked increase in Ci (by 25.1%) compared to control plants. application of HBL and was also effective (41.2%; 0.0265, p≤0.05) over unstressed control than their individual applications in enhancing E in control plants. application of EBL and HBL considerably increased the Fv/Fm and ФPSII in comparison to unstressed control. Control plants treated with EBL exhibited the impact on RuBPcase activity (43.1%; 0.0411, p≤0.05) over stressed control than their individual applications over the control plants. application of EBL was more effective (66.7%; 0.0237, p≤0.05) over unstressed control than their individual applications in enhancing FBPase activity in control plants. Control plants treated with EBL and HBL was found to be less effective on the PGK activity by 66.2% (0.0112, p≤0.05) than their individuals (EBL by 163.9%) compared to control. EBL and HBL has a more significant effect than their individual applications on the improvement of leaf starch and sucrose concentrations in drought stressed and well-watered plants.