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Query: EC:3.1.3.9 (glucose-6-phosphatase)
 3,081 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

L-Proline and L-glutamine were used to probe the inverse relationship between glycogenesis and ureagenesis in isolated, perfused livers from 48-h fasted rats. Both amino acids may provide nitrogen in the form of NH+4 for carbamyl-P synthesis. However, one molecule of glutamine may provide additionally for the synthesis of one molecule of the urea cycle substrate L-aspartate, but proline can provide for the synthesis of a molecule of NH+4 or one molecule of aspartate on an either/or basis only. In all perfusates, glucose was initially 30 mM (to favor phosphotransferase activity of glucose-6-phosphatase) and 0.5 mM 3-mercaptopicolinate was present (to inhibit glyconeogenesis from endogenous substrates, from the added amino acids, and via the indirect pathway). Glycogenesis from glucose, perfusate and hepatic urea formation, and levels of hepatic glucose-6-P, citrulline, PPi, and carbamyl-P were measured. The addition of glutamine to the perfusate markedly stimulated the urea cycle, but not glycogenesis. Hepatic urea level, perfusate urea concentration, and hepatic citrulline and PPi increased while carbamyl-P content decreased. In contrast, proline stimulated glycogenesis from glucose, but not ureagenesis. In the proline-supplemented compared with glutamine group, hepatic glycogenesis and carbamyl-P content increased; hepatic glucose-6-P levels showed a tendency toward increase; and hepatic urea formation, hepatic citrulline, and PPi levels were decreased. These observations are interpreted to support an hepatic mechanism whereby the relative availability of carbamyl-P to the urea cycle and as a substrate for glucose phosphorylation via phosphotransferase activity of the glucose-6-phosphatase system preliminary to glycogenesis from glucose is a major metabolic determinant.
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PMID:Reciprocal effects of proline and glutamine on glycogenesis from glucose and ureagenesis in isolated, perfused rat livers. 834 17

To examine the effects of the presence of Ehrlich ascites tumours on both the catalytic unit and the substrate/product translocase components of the glucose-6-phosphatase system in vivo, we isolated microsomes from the livers of control and tumour-bearing mice. Samples were analysed immunochemically for the quantity of catalytic unit, stabilizing protein and translocases T2 and T3 proteins. In comparison experiments, a variety of kinetic studies were performed. The most striking findings in tumour-bearing mice were: a 2.5-fold increase in the quantity of translocase T2 protein; increases in the Km and Vmax. for glucose 6-phosphate phosphohydrolase; and a decrease in the Km value for carbamoyl phosphate (carbamoyl-P) of carbamoyl-P:glucose phosphotransferase, all with intact microsomes. The percentage latency at Vmax. decreased for PPi phosphohydrolase and for glucose 6-phosphate phosphohydrolase, but was unaffected for carbamoyl-P:glucose phosphotransferase. These observations support a tumour-related increase in translocase T2 capacity in vivo, as it transports Pi from the microsomal lumen to the medium and carbamoyl-P or PPi from the medium to the microsomal lumen.
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PMID:The hepatic glucose-6-phosphatase system in Ehrlich-ascites-tumour-bearing mice. 838 51

There now is compelling evidence that hydrolysis of glucose-6-phosphate (Glc-6-P) in intact hepatic endoplasmic reticulum (ER) membrane preparations involves four integral components of the membrane: a Glc-6-P specific transporter (T1), a nonspecific enzyme (E) with its active site facing the lumen, and two other transport systems to mediate rapid and reversible fluxes of the hydrolytic products, inorganic phosphate (Pi) and glucose, i.e. (T2) and (T3), respectively. T2 also mediates transport of inorganic pyrophosphate (PPi) and carbamylphosphate. This concept readily and completely reconciles all known characteristics of the glucose-6-phosphatase (Glc-6-P'ase) system provided appropriate considerations are given to: (1) the quantitative contribution of E residing in membranes lacking a permeability barrier; (2) the kinetic restrictions imposed by T1 and T2; and (3) the influences of the endocrine, developmental and nutritional state on the kinetic relationship between the capacities to transport and hydrolyze. A broader-based understanding and application of these principles in the study of Glc-6-P'ase is needed to ensure accurate diagnosis of type 1 glycogen storage disease (GSD) and minimize unnecessary controversy. The view that the enzyme in native ER membranes is conformationally constrained is not supported by direct measurements of the catalytic turnover number. Finally, we describe the marked deficiencies of rapid filtration assays of Glc-6-P and PPi "uptake" as a direct method of diagnosis of types 1b and 1c GSD.
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PMID:Glucose-6-phosphatase and type 1 glycogen storage disease: some critical considerations. 839 48

Glucose-6-phosphatase is a multicomponent endoplasmic reticulum system comprising at least six different proteins, including a lumenal enzyme and several transport proteins. One of the transport proteins, T2beta, transports the substrate pyrophosphate and the product phosphate and its genetic deficiency is termed type 1c glycogen storage disease. We have used anti-T2beta antibodies for immunohistochemistry with image analysis and kinetic analysis of the glucose-6-phosphatase system to study for the temporal and spatial development of T2beta in human embryonic and fetal kidney. In metanephric kidney, there is an early predominance of T2beta expression in the ureteric bud derivatives and this changes with ontogeny such that developing nephrons, particularly proximal tubules, become dominant by mid-gestation. T2beta has the same spatial and temporal pattern as the glucose-6-phosphatase enzyme in both mesonephric and metanephric kidney. Pyrophosphate transport capacity is appropriate for the amount of glucose-6-phosphatase activity present in mid-gestation fetal kidney, in contrast to liver, where pyrophosphate transport capacity is developmentally delayed. Increasing knowledge of the temporal and spatial expression of the glucose-6-phosphatase proteins and their catalytic roles in early human development is essential for the elucidation of the aetiology of renal disease in both type I glycogen storage diseases and the developmental disorders of the glucose-6-phosphatase system.
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PMID:The human embryonic-fetal kidney endoplasmic reticulum phosphate-pyrophosphate transport protein. 860 68

N-Bromoacetylethanolamine phosphate (BAEP) has been used previously as an affinity label to study the hexose phosphate binding sites of fructose-6-P, 2-kinase:fructose-2, 6-bisphosphatase (Sakakibara et al. (1984) J. Biol. Chem. 259, 14023-14028). We have employed this compound to probe components of the glucose-6-phosphatase system using a combination of time-dependent and immediate inhibition kinetic techniques. Inhibition of D-glucose-6-phosphate (G6P) phosphohydrolase activity of native microsomes was irreversible and time- and inhibitor-concentration-dependent. Only a partial time-dependent, irreversible inhibition of the PPi phosphohydrolase activity of native microsomes was observed. BAEP inhibited PPi:glucose phosphotransferase activity of native microsomes in a concentration-dependent, irreversible manner which was more extensive than that seen with PPi phosphohydrolase, but less extensive than was observed with G6P phosphohydrolase. Disruption of microsomal integrity by detergent-treatment either prior to incubation with BAEP or subsequent to preliminary incubation with BAEP but prior to assay for activity abolished the time-dependent inhibition. These irreversible, time- and concentration-dependent inhibitory actions of BAEP thus are manifest at a site or sites where the intact membrane-bound enzyme first makes contact with substrates G6P and PPi. An additional site of inhibition by BAEP, through relatively weak, reversible competitive inhibition at the active catalytic site, is indicated by classical steady-state kinetic analysis. The irreversible, time- and concentration-dependent inhibitions by BAEP seen with G6P and PPi as substrates strongly suggest the potential utility of radio-labeled BAEP as an affinity label for the identification and ultimate isolation and study of uncharacterized auxiliary components of the glucose-6-phosphatase system.
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PMID:Inhibition of the glucose-6-phosphatase system by N-bromoacetylethanolamine phosphate, a potential affinity label for auxiliary proteins. 891 28

S 5627 is a synthetic analogue of chlorogenic acid. S 5627 is a potent linear competitive inhibitor of glucose 6-phosphate (Glc-6-P) hydrolysis by intact microsomes (Ki = 41 nM) but is without effect on the enzyme in detergent- or NH4OH-disrupted microsomes. 3H-S 5627 was synthesized and used as a ligand in binding studies directed at characterizing T1, the Glc-6-P transporter. Binding was evaluated using Ca2+-aggregated microsomes, which can be sedimented at low g forces. Aside from a modest reduction in K values for both substrate and S 5627, Ca2+ aggregation had no effect on glucose-6-phosphatase (Glc-6-Pase). Scatchard plots of binding data are readily fit to a simple "two-site" model, with Kd = 21 nM for the high affinity site and Kd = 2 microM for the low affinity site. Binding to the high affinity site was competitively blocked by Glc-6-P (Ki = 9 microM), whereas binding was unaffected by mannose-6-phosphate, Pi, and PPi and only modestly depressed by 2-deoxy-D-glucose 6-phosphate, a poor substrate for Glc-6-Pase in intact microsomes. Thus the high affinity 3H-S 5627 binding site fits the criteria for T1. Permeabilization of the membrane with 0.3% (3-[(chloramidopropyl)-dimethylammonio]-1-propanesulfonate) activated Glc-6-Pase and broadened its substrate specificity, but it did not significantly alter the binding of 3H-S 5627 to the high affinity sites or the ability of Glc-6-P to block binding. These data demonstrate unequivocally that two independent Glc-6-P binding sites are involved in the hydrolysis of Glc-6-P by intact microsomes. The present findings are the strongest and most direct evidence to date against the notion that the substrate specificity and the intrinsic activity of Glc-6-Pase in native membranes are determined by specific conformational constraints imposed on the enzyme protein. These data constitute compelling evidence for the role of T1 in Glc-6-Pase activity.
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PMID:Direct evidence for the involvement of two glucose 6-phosphate-binding sites in the glucose-6-phosphatase activity of intact liver microsomes. Characterization of T1, the microsomal glucose 6-phosphate transport protein by a direct binding assay. 949 46

1. The hepatic glucose-6-phosphatase (G-6-Pase) kinetic variables from chickens were studied in intact and disrupted microsomes using two substrates: glucose-6-phosphate (G-6-P) and pyrophosphate (PPi). They were studied from embryonic life to 51 d of age. 2. The phosphohydrolase activity studied in the broiler chicken liver microsomes corresponds to a true glucose-6-phosphatase. 3. The enzyme VMAX with both substrates in intact and disrupted microsomes showed 2 maxima: one in 19-d-old embryos and the other in 9-d-old chickens. Pyrophosphatase (PPase) VMAX in intact microsomes was higher than that of the G-6-Pase at all ages studied, except in 12 d embryos and 3-d-old chicks. In disrupted microsomes the VMAX of both enzymatic activities were similar. The G-6-Pase latency was high in the 19-d-old embryos and 51-d-old chickens. 4. The KM for PPi and G-6-Pase decreased when microsomes were disrupted. In intact microsomes the G-6-P KM was low in embryos and 3-d-old chicks and later increased. On the other hand, the PPi KM in intact microsomes showed little change during the animal's life and was lower than that of G-6-P. In disrupted microsomes the KM for both substrates were similar. 5. These results suggest a sequential incorporation of the G-6-Pase system components in the endoplasmic reticulum.
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PMID:Characterisation of hepatic microsomal glucose-6-phosphatase from broiler chickens. 1133 60


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