scholarly journals Glucose-6-Phosphate Dehydrogenase Regulation in Anoxia Tolerance of the Freshwater Crayfish Orconectes virilis

2011 ◽  
Vol 2011 ◽  
pp. 1-8 ◽  
Author(s):  
Benjamin Lant ◽  
Kenneth B. Storey
Keyword(s):  
Protein Kinase ◽  
Antioxidant Defense ◽  
Pentose Phosphate ◽  
Kinetic Properties ◽  
Freshwater Crayfish ◽  
Orconectes Virilis ◽  
Rate Determining Step ◽  
Flow Through ◽  
Glucose 6 Phosphate Dehydrogenase

Glucose-6-phosphate dehydrogenase (G6PDH), the enzyme which catalyzes the rate determining step of the pentose phosphate pathway (PPP), controls the production of nucleotide precursor molecules (R5P) and powerful reducing molecules (NADPH) that support multiple biosynthetic functions, including antioxidant defense. G6PDH from hepatopancreas of the freshwater crayfish (Orconectes virilis) showed distinct kinetic changes in response to 20 h anoxic exposure. Km values for both substrates decreased significantly in anoxic crayfish; Km NADP+ dropped from 0.015±0.008 mM to 0.012±0.008 mM, and Km G6P decreased from 0.13±0.02 mM to 0.08±0.007 mM. Two lines of evidence indicate that the mechanism involved is reversible phosphorylation. In vitro incubations that stimulated protein kinase or protein phosphatase action mimicked the effects on anoxia on Km values, whereas DEAE-Sephadex chromatography showed the presence of two enzyme forms (low- and high-phosphate) whose proportions changed during anoxia. Incubation studies implicated protein kinase A and G in mediating the anoxia-responsive changes in G6PDH kinetic properties. In addition, the amount of G6PDH protein (measured by immunoblotting) increased by ∼60% in anoxic hepatopancreas. Anoxia-induced phosphorylation of G6PDH could contribute to modifying carbon flow through the PPP under anoxic conditions, potentially maintaining NADPH supply for antioxidant defense during prolonged anoxia-induced hypometabolism.

scholarly journals Activity of key enzymes involved in glucose and triglyceride catabolism during bovine oocyte maturation in vitro

Reproduction ◽  
2002 ◽  
pp. 675-681 ◽  
Author(s):  
P Cetica ◽  
L Pintos ◽  
G Dalvit ◽  
M Beconi
Keyword(s):  
Enzymatic Activity ◽  
Pentose Phosphate Pathway ◽  
In Vitro Maturation ◽  
Specific Activity ◽  
Lipolytic Activity ◽  
Cumulus Cells ◽  
Pentose Phosphate ◽  
Key Enzymes ◽  
Glucose 6 Phosphate Dehydrogenase

Little is known about the metabolic profile of cumulus-oocyte complexes (COCs) during maturation. The aim of this study was to determine the differential participation of enzymatic activity in cumulus cells and the oocyte during in vitro maturation of bovine oocytes, by measuring the activity of key enzymes involved in the regulation of glycolysis (phosphofructokinase), the pentose phosphate pathway (glucose-6-phosphate dehydrogenase) and lipolysis (lipase). COCs were matured in medium 199 plus 10% (v/v) steer serum for 22-24 h at 39 degrees C in 5% CO(2):95% humidified air. Phosphofructokinase, glucose-6-phosphate dehydrogenase and lipase activities were measured in immature and in vitro matured COCs, denuded oocytes and cumulus cells, respectively. Phosphofructokinase and glucose-6-phosphate dehydrogenase activities (enzymatic units) remained constant during in vitro maturation of COCs, but there was a significant decrease in lipase activity (units) (P < 0.05), as activity in cumulus cells decreased significantly (P < 0.05). For the three enzymes studied, enzyme activity (units) remained unchanged in the oocyte during in vitro maturation. Specific activity increased in the oocyte (P < 0.05) and decreased in cumulus cells as a result of maturation (P < 0.05). In cumulus cells, phosphofructokinase was the most abundant of the three enzymes followed by glucose-6-phosphate dehydrogenase and then lipase (P < 0.05), whereas in the denuded oocyte this order was reversed (P < 0.05). Thus, the metabolism of cumulus cells is adapted to control the flow of metabolites toward the oocyte, which maintains its enzymatic activity even when dissociated from cumulus cells during maturation. The high activity of phosphofructokinase in cumulus cells indicates that glucose is metabolized mainly via the glycolytic pathway in these cells. The greater relative activity of glucose-6-phosphate dehydrogenase recorded in the oocyte indicates that glucose uptake could be directed mainly toward the pentose phosphate pathway. The marked lipolytic activity concentrated in the oocyte indicates an active participation in lipid catabolism during maturation.

scholarly journals Immunometabolic Endothelial Phenotypes: Integrating Inflammation and Glucose Metabolism

2021 ◽  
Author(s):  
Wusheng Xiao ◽  
William M Oldham ◽  
Carnen Priolo ◽  
Arvind K Pandey ◽  
Joseph Loscalzo
Keyword(s):  
Inflammatory Mediators ◽  
Nuclear Factor Κb ◽  
Pentose Phosphate ◽  
Pyruvate Dehydrogenase Kinase ◽  
Extracellular Flux ◽  
Pharmacological Blockade ◽  
Factor Α ◽  
Glucose 6 Phosphate Dehydrogenase

Rationale: Specific mechanisms linking inflammation and metabolic re-programming, two hallmarks of many pathobiological processes, remain incompletely defined. Objective: To delineate the integrative regulatory actions governing inflammation and metabolism in endothelial cells (ECs). Methods and Results: Metabolomic profiling, glucose labeling and tracing, and Seahorse extracellular flux analyses revealed that the inflammatory mediators, tumor necrosis factor α (TNFα) and lipopolysaccharide (LPS), extensively reprogram cellular metabolism, and particularly enhance glycolysis, mitochondrial oxidative phosphorylation (OXPHOS), and the pentose phosphate pathway (PPP) in primary human arterial ECs. Mechanistically, the enhancement in glycolysis and PPP is mediated by activation of the nuclear factor-κB (NF-κB)-6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase 3 (PFKFB3) axis and upregulation of glucose 6-phosphate dehydrogenase (G6PD), respectively; while enhanced OXPHOS was attributed to suppression of the forkhead box O1 (FOXO1)-pyruvate dehydrogenase kinase 4 (PDK4) axis. Restoration of the FOXO1-PDK4 axis attenuated the TNFα- or LPS-induced increase in OXPHOS but worsened inflammation in vitro, whereas enhancement of OXPHOS by pharmacological blockade of PDKs attenuated inflammation in mesenteric vessels of LPS-treated mice. Notably, suppression of G6PD expression or its activity potentiated the metabolic shift to glycolysis and/or endothelial inflammation, while inhibition of the NF-κB-PFKFB3 signaling, conversely, blunted the increased glycolysis and/or inflammation in in vitro and in vivo sepsis models. Conclusions: These results indicate that inflammatory mediators modulate the metabolic fates of glucose, and that stimulation of glycolysis promotes inflammation, whereas enhancement of OXPHOS and the PPP suppresses inflammation in the endothelium. Characterization of these immunometabolic phenotypes may have implications for the pathogenesis and treatment of many cardiovascular diseases.

The Compability with Enzyme Activity of Unusual Organic Osmolytes from Mangrove Red Algae

1996 ◽  
Vol 23 (5) ◽  
pp. 577 ◽  
Author(s):  
U Karsten ◽  
KD Barrow ◽  
O Nixdorf ◽  
RJ King
Keyword(s):  
Red Algae ◽  
Citric Acid Cycle ◽  
Compatible Solutes ◽  
Pentose Phosphate ◽  
Organic Osmolytes ◽  
Maximum Solubility ◽  
Organic Osmolyte ◽  
Negative Effect ◽  
Glucose 6 Phosphate Dehydrogenase

The effects of organic osmolytes synthesised and accumulated by red algae from mangrove habitats were investigated on the in vitro activities of two major enzymes, one of the citric acid cycle (malate dehydrogenase, MDH) and one of the oxidative pentose phosphate pathway (glucose-6- phosphate dehydrogenase, G6PDH). These enzymes were extracted from the mangrove algae Bostrychia tenella, Caloglossa leprieurii, Catenella nipae and Stictosiphonia hookeri. In each case, activity of the enzymes was inhibited with increasing NaCl concentrations up to 600 mM . In contrast, equimolar concentrations of mannitol (the major osmolyte in C. leprieurii), sorbitol (the major osmolyte in B. Tenella and S. hookeri) and a heteroside mixture (of which floridoside is the major osmolyte in C. nipae) did not inhibit enzyme function. Dulcitol, the second most important organic osmolyte in B. tenella, exerted no negative effect at its maximum solubility of 180 rnM on the salt-sensitive MDH. These data are all consistent with the proposed function of these organic compounds as compatible solutes.

scholarly journals Effect of adrenaline and phorbol myristate acetate or bacterial lipopolysaccharide on stimulation of pathways of macrophage glucose, glutamine and O2 metabolism. Evidence for cyclic AMP-dependent protein kinase mediated inhibition of glucose-6-phosphate dehydrogenase and activation of NADP+-dependent ‘malic’ enzyme

1995 ◽  
Vol 310 (2) ◽  
pp. 709-714 ◽  
Author(s):  
L F B P Costa Rosa ◽  
R Curi ◽  
C Murphy ◽  
P Newsholme
Keyword(s):  
Protein Kinase ◽  
Cyclic Amp ◽  
Pentose Phosphate Pathway ◽  
Malic Enzyme ◽  
Pentose Phosphate ◽  
Glutamine Metabolism ◽  
Dependent Protein Kinase ◽  
Macrophage Activity ◽  
Dependent Protein ◽  
Glucose 6 Phosphate Dehydrogenase

Adrenaline has recently been shown to stimulate both glucose metabolism and H2O2 release by macrophages but the activity of the key pentose phosphate pathway enzyme, glucose-6-phosphate dehydrogenase (which generates the NADPH crucial for the reduction of molecular oxygen), was reduced under these conditions [Costa Rosa, Safi, Cury and Curi (1992) Biochem. Pharmacol. 44, 2235-2241]. We report here that adrenaline activates another NADPH-producing enzyme, NADP(+)-dependent ‘malic’ enzyme, while also inhibiting glucose-6-phosphate dehydrogenase, via cyclic AMP-dependent protein kinase (PKA) activation. Regulation of glucose-6-phosphate dehydrogenase activity by PKA has not been reported elsewhere. The sparing of some glucose from pentose phosphate pathway consumption may be important in the provision of glycerol 3-phosphate which in the macrophage may be required for new phospholipid synthesis. Glutamine oxidation was also stimulated by adrenaline thus providing increased substrate (malate) for NADP(+)-dependent ‘malic’ enzyme and therefore shifting some of the burden of NADPH production from glucose to glutamine metabolism. We also report a novel synergistic effect of adrenaline and some bacterial products and/or gamma-interferon in stimulating secretory and metabolic pathways in macrophages which may be a part of a larger network of signals that lead to enhanced macrophage activity.

Inactivation of the AMP-activated protein kinase by glucose in cardiac myocytes: a role for the pentose phosphate pathway

2011 ◽  
Vol 32 (3) ◽  
pp. 229-239 ◽  
Author(s):  
Ikhlass Tabidi ◽  
David Saggerson
Keyword(s):  
Protein Kinase ◽  
Cardiac Myocytes ◽  
Pentose Phosphate Pathway ◽  
Adenine Nucleotide ◽  
Pentose Phosphate ◽  
Glucose Effect ◽  
Adult Rat ◽  
Ampk Activity ◽  
Amp Activated Protein Kinase ◽  
Glucose 6 Phosphate Dehydrogenase

Incubation of adult rat cardiac myocytes with increasing glucose concentrations decreased phosphorylation (αThr172) and activity of AMPK (AMP-activated protein kinase). The effect could be demonstrated without measurable changes in adenine nucleotide contents. The glucose effect was additive to the decrease in AMPK activity caused by insulin, was attenuated by adrenaline, was not mimicked by glucose analogues, lactate or pyruvate and was not due to changes in myocyte glycogen content. AMPK activity was decreased by xylitol and PMS (phenazine methosulfate) and was increased by the glucose-6-phosphate dehydrogenase inhibitor DHEA (dehydroepiandrosterone) and by thiamine. PMS and DHEA respectively, increased and decreased CO2 formation by the PPP (pentose phosphate pathway). AMPK activity was inversely related to the myocyte content of Xu5P (xylulose 5-phosphate), an intermediate of the non-oxidative arm of the PPP. Endothall, an inhibitor of PP2A (protein phosphatase 2A), abolished the glucose effect on AMPK activity. Further studies are needed to define the ‘active component’ that mediates the glucose effect and whether its site of action is PP2A.

scholarly journals Inhibition of the pentose phosphate shunt by lead: a potential mechanism for hemolysis in lead poisoning

Blood ◽  
1984 ◽  
Vol 63 (3) ◽  
pp. 518-524 ◽  
Author(s):  
NA Lachant ◽  
A Tomoda ◽  
KR Tanaka
Keyword(s):  
Lead Poisoning ◽  
Lead Acetate ◽  
Nicotinamide Adenine Dinucleotide Phosphate ◽  
Pentose Phosphate ◽  
Mechanism Of Inhibition ◽  
Glucose 6 Phosphate Dehydrogenase ◽  
Oxidant Sensitivity ◽  
Clinical Lead Poisoning ◽  
Pentose Phosphate Shunt

Abstract Recent investigations have disclosed a decrease in pentose phosphate shunt activity in hereditary pyrimidine 5′-nucleotidase deficiency. Clinical lead poisoning is associated with an acquired decrease in pyrimidine 5′-nucleotidase activity. The current investigations were undertaken (1) to determine if pentose shunt activity was decreased in erythrocytes exposed to lead, and (2) to compare the mechanism of inhibition to that seen in hereditary pyrimidine 5′-nucleotidase deficiency. Normal erythrocytes incubated with lead acetate in vitro demonstrated increased Heinz body formation, decreased reduced glutathione, a positive ascorbate cyanide test, and a reversible suppression of pentose shunt activity in the intact erythrocyte. Lead acetate added to normal red cell hemolysates markedly inhibited the activities of glucose-6-phosphate dehydrogenase (G6PD) and phosphofructokinase. The mean Kis of lead for glucose-6-phosphate and nicotinamide adenine dinucleotide phosphate (NADP) for G6PD were 1.5 microM and 2.1 microM, respectively, which is within the range of intraerythrocytic lead concentrations found in clinical lead poisoning. Magnesium enhanced the ability of lead to inhibit G6PD. Thus, the shortened erythrocyte survival in lead poisoning appears to be due, in part, to increased oxidant sensitivity secondary to inhibition of G6PD and the pentose shunt. The mechanism of shunt inhibition is, in part, similar to that seen in hereditary pyrimidine 5′-nucleotidase deficiency.

343 DISTRIBUTION OF GLUCOSE-6-PHOSPHATE DEHYDROGENASE AND IN VITRO MATURATION OF PORCINE OOCYTE - CUMULUS COMPLEXES FROM SMALL AND MEDIUM FOLLICLES

2007 ◽  
Vol 19 (1) ◽  
pp. 287
Author(s):  
Y. Ishida ◽  
H. Funahashi
Keyword(s):  
In Vitro Maturation ◽  
Critical Role ◽  
Morphological Characteristics ◽  
Cumulus Cells ◽  
Pentose Phosphate ◽  
G6pd Activity ◽  
Cresyl Blue ◽  
Key Enzyme ◽  
Glucose 6 Phosphate Dehydrogenase

Glucose metabolism through the pentose phosphate pathway (PPP) plays a critical role in meiotic maturation and fertilization. However, the relationship between the distribution of a PPP key enzyme, glucose-6-phosphate dehydrogenase (G6PD), in cumulus–oocyte complexes (COCs) and the in vitro maturation (IVM) of the oocytes is not clear. In the present study, we examined the distribution of G6PD, the morphological characteristics in OCCs derived from small (d2 mm in diameter) and medium (3 to 6 mm in diameter) follicles, and the rate of oocyte maturation. Porcine COCs were collected from small or medium follicles of slaughterhouse ovaries. The COCs were cultured in a maturation medium, BSA-free NCSU37 supplemented with 10% porcine follicular fluid, eCG, and hCG, for 20 h and then in the absence of hormones for 24 h. To determine the distribution of G6PD, the COCs were treated with 13 �M brilliant cresyl blue (BCB) in TL-HEPES-PVA for 90 min. Results from 3–6 replicates were analyzed by ANOVA and Duncan&apos;s multiple range test. The mean diameters for COCs collected from small follicles (136.7 &micro;m for the outer zona and 103.1 &micro;m for ooplasm) were significantly less than for those derived from medium follicles (164.1 &micro;m and 122.0 &micro;m, respectively). G6PD activity was detected in the cumulus cells for most of the COCs derived from medium follicles, but it was not significantly different from that of COCs derived from small follicles. In the second group of COCs, very little G6PD activity was found in both the cumulus cells and the oocytes (34.7 &plusmn; 11.5&percnt; and 18.0 &plusmn; 6.7&percnt; in COCS derived from small and medium follicles, respectively). After stimulation by eCG and hCG, the percentages of COCS in which G6PD activity was detected in the cumulus cells, but not in the oocytes, were 56.2 &plusmn; 23.8&percnt; and 72.9 &plusmn; 6.1&percnt; for small and medium follicles, respectively. The percentage of oocytes at the metaphase II stage (53&percnt; and 63.9&percnt; in oocytes from small and medium follicles, respectively) was higher for the COCs that showed higher G6PD activity in their cumulus cells. In conclusion, although no statistical differences were detected in the distribution of G6PD between COCs from small and medium follicles, due to a large variation, a higher percentage of mature oocytes seems to be the result of COCs where the G6PD activity is detected in the cumulus cells, but not in the oocyte, during IVM.

Inhibition of the pentose phosphate shunt by lead: a potential mechanism for hemolysis in lead poisoning

Blood ◽  
1984 ◽  
Vol 63 (3) ◽  
pp. 518-524
Author(s):  
NA Lachant ◽  
A Tomoda ◽  
KR Tanaka
Keyword(s):  
Lead Poisoning ◽  
Lead Acetate ◽  
Nicotinamide Adenine Dinucleotide Phosphate ◽  
Pentose Phosphate ◽  
Mechanism Of Inhibition ◽  
Glucose 6 Phosphate Dehydrogenase ◽  
Oxidant Sensitivity ◽  
Clinical Lead Poisoning ◽  
Pentose Phosphate Shunt

Recent investigations have disclosed a decrease in pentose phosphate shunt activity in hereditary pyrimidine 5′-nucleotidase deficiency. Clinical lead poisoning is associated with an acquired decrease in pyrimidine 5′-nucleotidase activity. The current investigations were undertaken (1) to determine if pentose shunt activity was decreased in erythrocytes exposed to lead, and (2) to compare the mechanism of inhibition to that seen in hereditary pyrimidine 5′-nucleotidase deficiency. Normal erythrocytes incubated with lead acetate in vitro demonstrated increased Heinz body formation, decreased reduced glutathione, a positive ascorbate cyanide test, and a reversible suppression of pentose shunt activity in the intact erythrocyte. Lead acetate added to normal red cell hemolysates markedly inhibited the activities of glucose-6-phosphate dehydrogenase (G6PD) and phosphofructokinase. The mean Kis of lead for glucose-6-phosphate and nicotinamide adenine dinucleotide phosphate (NADP) for G6PD were 1.5 microM and 2.1 microM, respectively, which is within the range of intraerythrocytic lead concentrations found in clinical lead poisoning. Magnesium enhanced the ability of lead to inhibit G6PD. Thus, the shortened erythrocyte survival in lead poisoning appears to be due, in part, to increased oxidant sensitivity secondary to inhibition of G6PD and the pentose shunt. The mechanism of shunt inhibition is, in part, similar to that seen in hereditary pyrimidine 5′-nucleotidase deficiency.

scholarly journals Phosphorylation of CK1δ: identification of Ser370 as the major phosphorylation site targeted by PKA in vitro and in vivo

2007 ◽  
Vol 406 (3) ◽  
pp. 389-398 ◽  
Author(s):  
Georgios Giamas ◽  
Heidrun Hirner ◽  
Levani Shoshiashvili ◽  
Arnhild Grothey ◽  
Susanne Gessert ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Phosphorylation Site ◽  
Fold Increase ◽  
Regulatory Subunit ◽  
Kinetic Properties ◽  
Casein Kinase 1 ◽  
Cell Extracts ◽  
Anchoring Protein

The involvement of CK1 (casein kinase 1) δ in the regulation of multiple cellular processes implies a tight regulation of its activity on many different levels. At the protein level, reversible phosphorylation plays an important role in modulating the activity of CK1δ. In the present study, we show that PKA (cAMP-dependent protein kinase), Akt (protein kinase B), CLK2 (CDC-like kinase 2) and PKC (protein kinase C) α all phosphorylate CK1δ. PKA was identified as the major cellular CK1δCK (CK1δ C-terminal-targeted protein kinase) for the phosphorylation of CK1δ in vitro and in vivo. This was implied by the following evidence: PKA was detectable in the CK1δCK peak fraction of fractionated MiaPaCa-2 cell extracts, PKA shared nearly identical kinetic properties with those of CK1δCK, and both PKA and CK1δCK phosphorylated CK1δ at Ser370in vitro. Furthermore, phosphorylation of CK1δ by PKA decreased substrate phosphorylation of CK1δ in vitro. Mutation of Ser370 to alanine increased the phosphorylation affinity of CK1δ for β-casein and the GST (gluthatione S-transferase)–p53 1–64 fusion protein in vitro and enhanced the formation of an ectopic dorsal axis during Xenopus laevis development. Anchoring of PKA and CK1δ to centrosomes was mediated by AKAP (A-kinase-anchoring protein) 450. Interestingly, pre-incubation of MiaPaCa-2 cells with the synthetic peptide St-Ht31, which prevents binding between AKAP450 and the regulatory subunit RII of PKA, resulted in a 6-fold increase in the activity of CK1δ. In summary, we conclude that PKA phosphorylates CK1δ, predominantly at Ser370in vitro and in vivo, and that site-specific phosphorylation of CK1δ by PKA plays an important role in modulating CK1δ-dependent processes.

scholarly journals Glucose 6-Phosphate Dehydrogenase from Trypanosomes: Selectivity for Steroids and Chemical Validation in Bloodstream Trypanosoma brucei

Molecules ◽  
2021 ◽  
Vol 26 (2) ◽  
pp. 358
Author(s):  
Cecilia Ortíz ◽  
Francesca Moraca ◽  
Marc Laverriere ◽  
Allan Jordan ◽  
Niall Hamilton ◽  
...  
Keyword(s):  
De Novo ◽  
Pentose Phosphate ◽  
Small Subset ◽  
Cell Physiology ◽  
Single Digit ◽  
In Vitro Proliferation ◽  
African Trypanosomes ◽  
Thiol Redox ◽  
Glucose 6 Phosphate Dehydrogenase

Glucose 6-phosphate dehydrogenase (G6PDH) fulfills an essential role in cell physiology by catalyzing the production of NADPH+ and of a precursor for the de novo synthesis of ribose 5-phosphate. In trypanosomatids, G6PDH is essential for in vitro proliferation, antioxidant defense and, thereby, drug resistance mechanisms. So far, 16α-brominated epiandrosterone represents the most potent hit targeting trypanosomal G6PDH. Here, we extended the investigations on this important drug target and its inhibition by using a small subset of androstane derivatives. In Trypanosoma cruzi, immunofluorescence revealed a cytoplasmic distribution of G6PDH and the absence of signal in major organelles. Cytochemical assays confirmed parasitic G6PDH as the molecular target of epiandrosterone. Structure-activity analysis for a set of new (dehydro)epiandrosterone derivatives revealed that bromination at position 16α of the cyclopentane moiety yielded more potent T. cruzi G6PDH inhibitors than the corresponding β-substituted analogues. For the 16α brominated compounds, the inclusion of an acetoxy group at position 3 either proved detrimental or enhanced the activity of the epiandrosterone or the dehydroepiandrosterone derivatives, respectively. Most derivatives presented single digit μM EC50 against infective T. brucei and the killing mechanism involved an early thiol-redox unbalance. This data suggests that infective African trypanosomes lack efficient NADPH+-synthesizing pathways, beyond the Pentose Phosphate, to maintain thiol-redox homeostasis.

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