Glucose 6 phosphate dehydrogenase

Manufactured by Roche

Sourced in Germany, Switzerland, Argentina, Japan

Glucose-6-phosphate dehydrogenase is an enzyme that catalyzes the first step of the pentose phosphate pathway, converting glucose-6-phosphate to 6-phosphoglucono-δ-lactone. The enzyme plays a crucial role in cellular metabolism and redox homeostasis.

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Lab products found in correlation

Hexokinase, by Roche (5 mentions) Glucose 6 phosphate (g6p), by Roche (4 mentions) Glucose 6 phosphate (g6p), by Merck Group (3 mentions) Bovine serum albumin (bsa), by Merck Group (2 mentions) Lactate dehydrogenase, by Roche (2 mentions) Nadph, by Merck Group (2 mentions) Hexokinase, by Oriental Yeast (2 mentions) Phosphoglucoisomerase, by Merck Group (2 mentions) Hexokinase, by Merck Group (2 mentions) Hepes, by Merck Group (1 mentions) Nifedipine, by Merck Group (1 mentions) Glibenclamide, by Merck Group (1 mentions) Dl dithiothreitol, by Merck Group (1 mentions) Collagenase p, by Roche (1 mentions) 6 propyl 2 thiouracil, by Merck Group (1 mentions) Dionex ics 3000 system, by Thermo Fisher Scientific (1 mentions) Nigerose, by Biosynth (1 mentions) Kojibiose, by Biosynth (1 mentions) Turanose, by Biosynth (1 mentions) Folin ciocalteu reagent, by Merck Group (1 mentions)

Close spelling variants of this product

Hexokinase/glucose-6-phosphate dehydrogenase assay (5 citations) Hexokinase/glucose-6-phosphate dehydrogenase enzymatic in vitro test (2 citations)

26 protocols using glucose 6 phosphate dehydrogenase

Preparation and Use of Chemical Reagents

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Nifedipine, glibenclamide, acetylcholine, sodium pentobarbital, bovine serum albumin (BSA), HEPES, DL-Dithiothreitol (DTT), and 6-propyl-2-thiouracil (PTU) were purchased from Sigma (St. Louis, MO, USA); collagenase P, NADP disodium salt, ATP disodium salt, glucose-6-phosphate dehydrogenase from Roche (Roche Diagnostic, Mannheim, Germany), and all other reagent-grade chemicals from Merck (Darmstadt, Germany).
Stock solutions of Nifedipine and glibenclamide, were prepared in ethanol and dimethyl sulphoxide (DMSO) and the other substances were dissolved in H₂O or directly added into the incubation media.

Godini A., Ghasemi A, & Zahediasl S. (2015). The Possible Mechanisms of the Impaired Insulin Secretion in Hypothyroid Rats. PLoS ONE, 10(7), e0131198.

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Comprehensive Biochemical Assay Protocol

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Acetic acid (PubChem CID:176), aminopyrine (PubChem CID:6009), bovine serum albumin (PubChem CID:16132389), dichlorophenolindophenol (PubChem CID:13726) (DCPIP), epinephrine (PubChem CID:5816), ethoxycoumarin (PubChem CID:35703), Folin-Ciocalteu reagent, glycerol (PubChem CID:753) from Sigma-Aldrich, glucose 6-phosphate (PubChem CID:5958) and glucose 6-phosphate dehydrogenase from Roche Diagnostic; L-glutathione oxidized (PubChem CID:71308714), L-glutathione reduced (PubChem CID:745), methanol (PubChem CID:5958), methoxyresorufin (PubChem CID:119220), Nicotinamide adenine dinucleotide phosphate in oxidized (PubChem CID:5886) and reduced form (PubChem CID:5886) (NADP+ and NADPH), p-nitrophenol (PubChem CID:980), pentoxyresorufin (PubChem CID:107683), perchloric acid (PubChem CID:24247), resorufin (PubChem CID:69462), sodium dithionite (PubChem CID:24489), trichloroacetic acid (PubChem CID:6421), Triton X-100 (PubChem CID:5590), Trizma (PubChem CID:16218782), umbelliferone (PubChem CID:4412127), 1chloro-2,4-dinitrobenzene (PubChem CID:6), 1-naphthol (PubChem CID:7005), 7-ethoxyresorufin (PubChem CID:3294) from Sigma-Aldrich. Vitamin E in the form of DL-all-rac α-Tocopherol (PubChem CID: 2116).
All others chemicals were highest purity commercially available.

Vivarelli F., Canistro D., Franchi P., Sapone A., Vornoli A., Della Croce C., Longo V., Lucarini M, & Paolini M. (2016). Disruption of redox homeostasis and carcinogen metabolizing enzymes changes by administration of vitamin E to rats. Life sciences, 145.

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Spectrophotometric Assessment of NQO1 Activity

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After the treatment of Hepa1c1c7 cells (1 × 10⁴ cells/well in 96-well plates), the activity of NQO1 was determined spectrophotometrically as described previously [21] (link). Cells were washed four times with PBS and lysed with 75 μL of digitonin solution (0.8 g/L digitonin, 2 mM EDTA, pH 7.8) by shaking on an orbital shaker for 20 min at room temperature. One part of the cell lysate (20 μL) was used to determine the protein content. The remaining lysate (55 μL) was mixed with 200 μL of 0.5 M Tris-Cl buffer containing 10% (w/v) bovine serum albumin, 1.5% (v/v) Tween-20, 7.5 mM FAD, 150 mM glucose-6-phosphate, 2 U/mL glucose-6-phosphate dehydrogenase (Roche Diagnostics, Mannheim, Germany), 50 mM NADP⁺, 25 mM menadione and 0.7 mM MTT. The mixture was incubated for 5 min at room temperature and the reaction was stopped with 50 μL of dicumarol suspension (0.3 mM dicumarol, 5 mM potassium phosphate, 0.5% DMSO). The absorbance of the reduced MTT corresponding to the activity of NQO1 was measured at 610 nm on a spectrophotometric plate reader. The absorbance values were normalized to the protein content and used for the calculation of fold changes versus the control.

Roubalová L., Dinkova-Kostova A.T., Biedermann D., Křen V., Ulrichová J, & Vrba J. (2017). Flavonolignan 2,3-dehydrosilydianin activates Nrf2 and upregulates NAD(P)H:quinone oxidoreductase 1 in Hepa1c1c7 cells. Fitoterapia, 119, 115-120.

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Enzymatic Assay of Rare Carbohydrates

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α-pNPG (N1377), β-pNPG (N7006), maltose (M5885), sucrose (S0389) and α-MG (M9376) were purchased from Sigma-Aldrich. Isomaltose (MI04560), palatinose (OI04225), isomaltotriose (OI05352), panose (OP06685), trehalose (OT03932), kojibiose (OK05039), nigerose (ON06975), turanose (OT06692), maltulose (OM06578), leucrose (OL06727), maltotriose (OM06486) and melezitose (OM00386) were purchased from Carbosynth. These sugars were chosen with the highest purity as possible and were further analysed by HPAE-PAD (Dionex ICS3000 system equipped with a carbopac PA10 analytical ion exchange column and pulsed amperometric detector) to confirm their purity. All other chemicals were purchased from Sigma. Glucose-6-phosphate dehydrogenase (101276550001) and hexokinase (11426362001) were purchased from Roche Diagnostics.

Deng X., Petitjean M., Teste M.A., Kooli W., Tranier S., François J.M, & Parrou J.L. (2014). Similarities and differences in the biochemical and enzymological properties of the four isomaltases from Saccharomyces cerevisiae. FEBS Open Bio, 4, 200-212.

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Spectrophotometric Enzymatic Assay Protocol

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Acetonitrile (PubChem CID:6342), aminopyrine (PubChem CID:6009), bovin serum albumin, dichlorophenolindophenol (PubChem CID:13726) (DCPIP), epinephrine (PubChem CID:5816), ethoxycoumarin (PubChem CID:35703), Folin-Ciocalteu reagent from Sigma-Aldrich, glusose 6-phosphate (PubChem CID:5958) and glucose 6-phosphate dehydrogenase from Roche Diagnostic (Indianapolis, IN, USA), L-glutathione oxidized (PubChem CID:71308714), L-glutathione reduced (PubChem CID:745), methanol (PubChem CID:5958) were HPLC grade, methoxyresorufin (PubChem CID:119220), Nicotinamide adenine dinucleotide phosphate in oxidized (PubChem CID:5886) and reduced form (PubChem CID:5886) (NADP+ and NADPH), p-nitrophenol (PubChem CID:980), pentoxyresorufin (PubChem CID:107683), phenylmethylsulfonyl fluoride, resorufin (PubChem CID:69462), sodium dithionite (PubChem CID:24489), Triton X-100, 1-chloro-2,4-dinitrobenzene (PubChem CID:6), 1-naphtol, 7-ethoxyresorufin (PubChem CID:3294) from Sigma-Aldrich.
All others chemicals were highest purity and are commercially available.

Vivarelli F., Canistro D., Sapone A., De Nicola G.R., Babot Marquillas C., Iori R., Antonazzo I.C., Gentilini F, & Paolini M. (2016). Raphanus sativus cv. Sango Sprout Juice Decreases Diet-Induced Obesity in Sprague Dawley Rats and Ameliorates Related Disorders. PLoS ONE, 11(3), e0150913.

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Measuring Lactate and Glucose in HAP1 Cells

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HAP1 cells (150 × 10³) were seeded in 12-well plates and incubated in IMDM with 10% FBS for 40 hr with a change of medium every 20 hr. Following 40 hr incubation, cell confluency was about 90% and the fresh medium was added to start the final incubation lasting for the next 20 hr. To finish the incubation, 1 ml of the culture medium was transferred to 100 μl of 35% HClO₄ and centrifuged at 10,000 × g for 10 min, before the resulting supernatant was neutralized with 3 M K₂CO₃/3 M KOH. The lactate produced was determined spectrophotometrically using lactate dehydrogenase (Roche) (Noll, 1984 ), whereas glucose consumption was measured in a spectrophotometric assay that uses hexokinase and glucose-6-phosphate dehydrogenase (Roche) (Kunst and Draeger, 1984 ). Adherent cells were washed three times with 3 ml of PBS and then solubilized in 0.1 M NaOH containing 0.125% Triton X-100 for protein quantification (Bradford) to allow normalization of lactate produced and glucose consumed to biomass.

Kwiatkowski S., Seliga A.K., Vertommen D., Terreri M., Ishikawa T., Grabowska I., Tiebe M., Teleman A.A., Jagielski A.K., Veiga-da-Cunha M, & Drozak J. (2018). SETD3 protein is the actin-specific histidine N-methyltransferase. eLife, 7, e37921.

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Cytochrome P450 Enzyme Assay Protocol

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Recombinant human acetylcholinesterase, superoxide dismutase, horseradish peroxidase, NADPH, menadione, acetylthiocholine chloride, 5,5′-dithiobis(2-nitrobenzoic acid), 7-ethoxyresorufin, tetraisopropyl pyrophosphoramide, and coumarin were purchased from Sigma (St. Louis, MO). Parathion, paraoxon, and diethylthiophosphate were from Chem Service Inc. (West Chester, PA). Amplex Red reagent was from Molecular Probes (Eugene, OR). Recombinant human CYPs (CYP1A1, CYP1A2, CYP1B1, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, CYP3A4, CYP3A5 and CYP 3A7), and pooled human liver microsomes were purchased from BD Gentest (Woburn, MA). Recombinant human CPR, Vivid P450 substrates, 7-ethoxy-methyloxy-3-cyanocoumarin (EOMCC) and 7-benzyloxy-methyloxy-3-cyanocoumarin (BOMCC), 7-methoxy-4-trifluoromethyl coumarin and dibenzylfluorescein were from Life Technologies (Grand Island, NY). Glucose-6-phosphate and Glucose-6-phosphate dehydrogenase were from Roche Diagnostic (Indianapolis, IN).

Jan Y.H., Richardson J.R., Baker A.B., Mishin V., Heck D.E., Laskin D.L, & Laskin J.D. (2015). Vitamin K3 (menadione) redox cycling inhibits cytochrome P450-mediated metabolism and inhibits parathion intoxication. Toxicology and applied pharmacology, 288(1), 114-120.

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Starch Quantification from Rosette Leaves

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Starch was extracted and assayed as previously described (Smith and Zeeman, 2006) . Frozen rosette leaves were ground to a fine powder and homogenized in 8% (v/v) perchloric acid (Ricca Chemical, Arlington, TX, USA). The insoluble fraction was pelleted and washed 3 times in 80% (v/v) ethanol and resuspended in water. Prior to quantification, the samples were boiled 15 min at 95°C to gelatinize the starch and hydrolyzed with α-amylase and amyloglucosidase (Roche, Basel, Switzerland) to release the glucose blocks. Glucose content was then measured by spectrophotometry after a two enzymes reaction with hexokinase and glucose-6-phosphate dehydrogenase (Roche).

Scandola S., Mehta D., Li Q., Rodriguez Gallo M.C., Castillo B, & Uhrig R.G. (2022). Multi-omic analysis shows REVEILLE clock genes are involved in carbohydrate metabolism and proteasome function. Plant physiology, 190(2).

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Starch Quantification via Enzymatic Digestion

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Samples were autoclaved for 20 min at 120 °C to solubilise the starch. Starch was digested to glucose using α-amylase and amyloglucosidase (both from Aspergillus oryzae, manufactured by Megazyme Ltd., Co. Wicklow, Ireland) and the glucose measured enzymatically as described in Rösti et al. (2006) (link). Coupling enzymes in the glucose assay, hexokinase and (NADP-dependant) glucose-6-phosphate dehydrogenase were manufactured by Roche Diagnostics (Basel, Switzerland). The reaction was monitored at 340 nm on either a spectrophotometer or a microtitre plate reader. Initial absorbance was recorded before the reaction was started by the addition of (NADP-dependant) glucose-6-phosphate dehydrogenase. The reaction was monitored until steady absorbance was reached. The difference between initial absorbance and final absorbance was used to calculate glucose in the samples. The mass of starch (g) was calculated as moles of glucose ×162. Undigested controls were included in the analysis to adjust for glucose in the pellet not from starch.

Cook F., Hughes A., Nibau C., Orman-Ligeza B., Schatlowski N., Uauy C, & Trafford K. (2018). Barley lys3 mutants are unique amongst shrunken-endosperm mutants in having abnormally large embryos. Journal of Cereal Science, 82, 16-24.

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Enzymatic Activity Assay Protocol

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All laboratory chemicals, substrates and pigeon liver acetone powder were purchased from Sigma Aldrich (St. Louis, MO), except for auxiliary enzymes glucose-6-phosphate dehydrogenase and lactate dehydrogenase (Roche Diagnostics, Indianapolis, IN), bovine serum albumin (MP Biochemicals, Santa Ana, CA), BioRad protein assay kits (BioRad, Hercules, CA) and the dye p-(p-aminophenylazo)-benzene sulphonic acid (Alfa Aesar, Ward Hill, MA).

Hagopian K., Tomilov A.A., Kim K., Cortopassi G.A, & Ramsey J.J. (2015). Key Glycolytic Enzyme Activities of Skeletal Muscle Are Decreased under Fed and Fasted States in Mice with Knocked Down Levels of Shc Proteins. PLoS ONE, 10(4), e0124204.

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