Oxidative Enzymes and Pathways of Hexose and Triose Metabolism in Chlorella

Cell-free preparations of Chlorella pyrenoidosa Chick, van Niel's strain, were assayed for oxidative enzymes, utilizing isotopic and spectrophotometric techniques. The enzyme activity of heterotrophic and autotrophic cells was compared. The study was divided into categories, one concerned with the spectrophotometric detection of enzymes involved in the initial reactions of glycolysis and the hexose monophosphate shunt, and the other with the direct oxidation of glucose as compared with that oxidized via glycolysis. The reduction of pyridine nucleotides in crude extracts was studied with glucose, glucose-6-phosphate, 6-phosphogluconate, and fructose-1-6-diphosphate as substrates. Enzymes detected in both heterotrophic and autotrophic cells were hexokinase, fructose-diphosphate-aldolase, NAD-linked 3-phosphoglyceraldchyde dehydrogenase, glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, and a NADP-linked 3-phosphoglyceraldchyde dehydrogenase. In addition to isotopic studies designed to make an appraisal of the hexose monophosphate shunt, a comparison of the rate of reduction of NADP by glucose-6-phosphate and 6-phosphogluconate in relation to the reduction of NAD by 3-phosphoglyceraldehyde was made in light- and dark-grown cells. The rate of reduction of NADP appeared to be lowered in the light-grown cells, suggesting, as did also the isotopic studies, that the hexose monophosphate shunt is less active in autotrophic metabolism than in heterotrophic metabolism.     

Cyclic organization of the carbohydrate metabolism in Sinorhizobium meliloti.

The pathways of polysaccharide biosynthesis were investigated in cells of Sinorhizobium meliloti (strain Su47) using a stable isotope approach. The isotopic labeling of the periplasmic beta-1,2-glucans synthesized from glucose labeled at various positions evidenced the involvement of catabolic pathways, namely the pentose-phosphate and Entner-Doudoroff pathways, into the early steps of polysaccharide synthesis. The exopolysaccharides produced at the same time had a labeling pattern similar to that of the beta-glucans, indicating similar early steps for both polysaccharides. The results emphasized a cyclic organization of the carbohydrate metabolism in S. meliloti, in which the carbons of the initial hexose were allowed to re-enter the catabolic pathways many times. The metabolic incidences of such metabolic topology are discussed.     

The effect of hypoxia on the control of carbohydrate metabolism in ripening bananas

The aim of this work was to determine the effects of hypoxia on the major fluxes of carbohydrate metabolism in climacteric fruit of banana (Musa cavendishii Lamb ex Paxton). Hands of bananas, untreated with ethylene, were allowed to ripen in air at 21°C in the dark. When the climacteric began, fruit were transferred to 15 or 10% oxygen and were analysed once the climacteric peak had been reached 8–12 h later. The rates of starch breakdown, sucrose, glucose and fructose accumulation, and CO₂ production were determined, as were the contents of hexose monophosphates, adenylates and pyruvate. In addition, the detailed distribution of label was determined after supplying [U-¹⁴C]-, [1-¹⁴C]-, [3,4-¹⁴C]- and [6-¹⁴C]glucose, and [U-¹⁴C]glycerol to cores of tissue under hypoxia. The data were used to estimate the major fluxes of carbohydrate metabolism. There was a reduction in the rate of respiration. The ATP/ADP ratio was unaffected but there was a significant increase in the content of AMP. In 15% oxygen only minor changes in fluxes were observed. In 10% oxygen starch breakdown was reduced and starch synthesis was not detected. The rate of sucrose synthesis decreased, as did the rate of re-entry of hexose sugars into the hexose monophosphate pool. There was a large increase in both the glycolytic flux and in the flux from triose phosphates to hexose monophosphates. It is argued that the increase in these fluxes is due to activation of pyrophosphate: fructose-6-phosphate 1-phosphotransferase, and that this enzyme has an important role in hypoxia. The results are discussed in relation to our understanding of the control of carbohydrate metabolism in hypoxia.Abbreviations Glc6P glucose-6-phosphate - Glc1P glucose-1-phosphate - Fru6P fructose-6-phosphate - PP_i inorganic pyro-phosphate We thank Geest Foods Group, Great Dunmow, Essex, UK for giving us the bananas. S.A.H. thanks the managers of the Brood bank Fund for a fellowship.     

Mutational Analyses of Glucose Dehydrogenase and Glucose-6-Phosphate Dehydrogenase Genes in Pseudomonas fluorescens Reveal Their Effects on Growth and Alginate Production

The biosynthesis of alginate has been studied extensively due to the importance of this polymer in medicine and industry. Alginate is synthesized from fructose-6-phosphate and thus competes with the central carbon metabolism for this metabolite. The alginate-producing bacterium Pseudomonas fluorescens relies on the Entner-Doudoroff and pentose phosphate pathways for glucose metabolism, and these pathways are also important for the metabolism of fructose and glycerol. In the present study, the impact of key carbohydrate metabolism enzymes on growth and alginate synthesis was investigated in P. fluorescens. Mutants defective in glucose-6-phosphate dehydrogenase isoenzymes (Zwf-1 and Zwf-2) or glucose dehydrogenase (Gcd) were evaluated using media containing glucose, fructose, or glycerol. Zwf-1 was shown to be the most important glucose-6-phosphate dehydrogenase for catabolism. Both Zwf enzymes preferred NADP as a coenzyme, although NAD was also accepted. Only Zwf-2 was active in the presence of 3 mM ATP, and then only with NADP as a coenzyme, indicating an anabolic role for this isoenzyme. Disruption of zwf-1 resulted in increased alginate production when glycerol was used as the carbon source, possibly due to decreased flux through the Entner-Doudoroff pathway rendering more fructose-6-phosphate available for alginate biosynthesis. In alginate-producing cells grown on glucose, disruption of gcd increased both cell numbers and alginate production levels, while this mutation had no positive effect on growth in a non-alginate-producing strain. A possible explanation is that alginate synthesis might function as a sink for surplus hexose phosphates that could otherwise be detrimental to the cell.     

Recycling of carbon in the oxidative pentose phosphate pathway in non-photosynthetic plastids

Recycling of carbon in the oxidative pentose phosphate pathway (OPPP) of intact pea root plastids has been studied. The synthesis of dihydroxyacetone phosphate (DHAP) and evolution of CO₂ was followed in relation to nitrite reduction. A close coupling was observed between all three measured fluxes which were linear for up to 60 min and dependent upon the integrity of the plastids. However, the quantitative relationship between 1-¹⁴CO₂ evolution from glucose 6-phosphate and nitrite reduction varied with available hexose phosphate concentration. When 10 mM glucose 6-phosphate was supplied to intact plastids a stoichiometry of 1.35 was observed between ¹⁴CO₂ evolution and nitrite reduction. As exogenous glucose 6-phosphate was decreased this value fell, becoming 0.47 in the presence of 0.2 mM glucose 6-phosphate, indicative of considerable recycling of carbon. This conclusion was reinforced when using [2-¹⁴C]glucose-6-phosphate. The measured release of 2-¹⁴CO₂ was consistent with the data for 1-¹⁴CO₂, suggesting complete recycling of carbon in the OPPP. Ribose 5-phosphate was also able to support nitrite reduction and DHAP production. A stoichiometry of 2 NO ₂ ^? reduced: 1 DHAP synthesised was observed at concentrations of 1 mM ribose 5-phosphate or less. At concentrations of ribose 5-phosphate greater than 1 mM this stoichiometry was lost as a result of enhanced DHAP synthesis without further increase in nitrite reduction. It is suggested that this decoupling from nitrite reduction is a function of excess substrate entering directly into the non-oxidative reactions of the OPPP, and may be useful when the demand for OPPP products is not linked to the demand for reductant. The significance of recycling in the OPPP is discussed in relation to the coordination of nitrate assimilation with carbohydrate oxidation in roots and with the utilisation of carbohydrate by other pathways within plastids.     

The response of carbohydrate metabolism in potato tubers to low temperature

This work investigates the possible causes of cold-induced sweetening in potato by examining the impact of low temperature on carbohydrate metabolism in mature tubers. Metabolism in tuber discs was monitored by determining the redistribution of radiolabel following incubation in [U-(14)C]glucose. Estimates of flux based on the specific activity of hexose phosphates established that while incubation at 4 degrees C resulted in an immediate restriction in pathways of carbohydrate oxidation relative to activity at 25 degrees C, there was no corresponding increase in flux to soluble sugars. In contrast, prior storage at low temperature stimulated flux to sugars at both 4 and 25 degrees C. Comparison of (14)CO(2) release from specifically labeled glucose and gluconate fed to tuber discs at 4 and 25 degrees C indicated that flux through glycolysis was preferentially restricted relative to the oxidative pentose phosphate pathway at low temperature, irrespective of prior storage temperature. However, the degree of randomization of label between positions C1 and C6 in the fructosyl moiety of sucrose following metabolism of [1-(13)C]glucose established that there was no preferential inhibition of the recycling of triose phosphates to hexose phosphates at low temperature. These results indicate that sugar accumulation in tubers during storage in the cold is not a direct consequence of a constraint in carbohydrate oxidation, despite preferential restriction of glycolysis at low temperature. It is concluded that the cold lability of enzymes catalyzing the conversion of fructose 6-phosphate to fructose 1,6-bisphosphate is not a major factor in cold-induced sweetening in plants and that this widely held hypothesis should be abandoned.     

Changes in the rates of synthesis and messenger RNA levels of hepatic glucose-6-phosphate and 6-phosphogluconate dehydrogenases following induction by diet or thyroid hormone

The lipogenic capacity of rat liver is increased in animals fed a high carbohydrate, fat-free diet or by the administration of 2,2',5'-triiodo-L-thyronine. Underlying this change is a generalized induction of the enzymes involved in lipogenesis, including glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, and malic enzyme, which together serve to generate the additional NADPH required for increased fatty acid synthesis. This report presents evidence indicating that induction of the hexose-shunt dehydrogenases involves increased enzyme synthesis secondary to elevated enzyme specific mRNA levels, as has previously been shown for malic enzyme. Activities of specific mRNAs, estimated by cell-free translation of hepatic poly(A)-containing RNA in the mRNA dependent rabbit reticulocyte lysate, were compared with enzyme specific activities and relative rates of specific enzyme synthesis. The 2-fold increase in glucose-6-phosphate dehydrogenase specific activity in hyperthyroid rats and the 13-fold increase in rats fed a high carbohydrate, fat-free diet, relative to euthyroid, chow-fed controls were paralleled by comparable increases in the synthetic rates and mRNA levels of this enzyme. Similarly, consonant changes in the rate of enzyme synthesis and concentration of 6-phosphogluconate dehydrogenase mRNA accompanied the 2.5- and 3-fold increases in specific activity of this enzyme observed in response to hormonal and dietary induction, respectively. Thus, both thyroid hormone and carbohydrate feeding appear to induce glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase primarily by increasing the effective cellular concentrations of their respective mRNAs and, consequently, their rates of synthesis.     

The effect of glucose on the control of carbohydrate metabolism in ripening bananas

The effect of exogenous glucose on the major fluxes of carbohydrate metabolism in cores of climacteric fruit of banana (Musa cavendishii Lamb ex Paxton) was determined with the intention of using the effects in the application of top-down metabolic control analysis. Hands of bananas, untreated with ethylene, were allowed to ripen in the dark at 21 °C. Cores were removed from climacteric fruit and incubated in 100 or 200 mM glucose for 4 or 6 h. The rates of starch breakdown, sucrose and fructose accumulation and CO₂ production were measured. The steady-state contents of hexose monophosphates, adenylates and pyruvate were determined. In addition, the detailed distribution of label was determined after supply of the following: [U-¹⁴C]-, [1-¹⁴C]-, [3,4¹⁴C]and [6-¹⁴C]glucose, and [U-¹⁴C]glycerol. The data were used to estimate the major fluxes of carbohydrate metabolism. Supply of exogenous glucose led to increases in the size of the hexose-monophosphate pools. There was a small stimulation of the rate of sugar synthesis and a major increase in the rate of starch synthesis. Starch breakdown was inhibited. Respiration responded to the demand for ATP by sugar synthesis. The effect of glucose on fluxes and metabolite pools is discussed in relation to our understanding of the control and regulation of carbohydrate metabolism in ripening fruit.Abbreviations Glc6P glucose-6-phosphate - Glc1P glucose-1-phosphate - Fru6P fructose-6-phosphate - AEC adenylate energy charge We thank Geest Foods Group, Great Dunmow, Essex, UK for giving us the bananas. SAH thanks the managers of the Broodbank Fund for a fellowship.     

Hexose metabolism in pancreatic islets: enzyme-to-enzyme tunnelling of hexose 6-phosphates.

The fate of unlabelled D-glucose and D-[2-3H]glucose in pancreatic islets was simulated taking into account experimental values for glycolytic flux, intracellular concentration of D-glucose 6-phosphate and phosphoglucoisomerase activity. The model, which also takes into account the isotopic discrimination in velocity and intramolecular transfer of tritium between D-[2-3H]glucose 6-phosphate and D-[1-3H]fructose 6-phosphate in the reaction catalyzed by phosphoglucoisomerase, revealed that the predicted generation of 3HOH from D-[2-3H]glucose was much higher than the true experimental value. Such a discrepancy is reinforced by the consideration that the generation of 3HOH from D-[2-3H]glucose in islet cells is not solely attributable to the phosphoglucoisomerase-catalyzed detritiation of hexose 6-phosphates metabolized in the glycolytic pathway. In order to reconcile experimental and theoretical values for 3HOH production, it was found necessary to postulate enzyme-to-enzyme tunnelling of hexose 6-phosphates in the hexokinase/phosphoglucoisomerase/phosphofructokinase sequence. It is proposed that such a tunnelling may favour the anomeric specificity of D-glucose metabolism in islet cells, by restricting the anomerization of hexose 6-phosphates.     

Compartmentation in the induction of the hexose-6-phosphate transport system of Escherichia coli

The induction of the hexose-6-phosphate transport system was investigated. Glucose-6-phosphate (G6P) at concentrations as low as 10(-4)m was able to induce this system in wild-type cells, as well as in mutants lacking phosphoglucose isomerase or G6P dehydrogenase. Growth in the presence of fructose-6-phosphate (F6P) induced the system only if the cells contained phosphoglucose isomerase. Furthermore, glucose and F6P were found to induce the system only if the extracellular concentration of G6P became appreciable in the medium as a consequence of the leakage of intracellular G6P formed from the glucose or F6P. Intracellular G6P was not an inducer even at high concentrations. The metabolism of glucose inhibited the induction of the hexose-6-phosphate transport system. Hypotheses for this compartmentalization of inducer and membrane-associated induction are presented.     

Phosphoglucoisomerase-catalyzed interconversion of hexose phosphates: a model for the interconversion of D-[2-3H]glucose 6-phosphate and D-[1-3H]fructose 6-phosphate

Based on experimental data, a model is proposed for the interconversion of either unlabelled hexose phosphates or D-[2-3H]glucose 6-phosphate and D-[1-3H]fructose 6-phosphate in the reaction catalyzed by phosphoglucoisomerase. This model takes into account the known differences in maximal velocity and affinity for each substrate, the intramolecular transfer of tritium between C1 and C2, and the isotopic discrimination between unlabelled and tritiated esters. This model reveals that, in a close system characterized by the progressive detritiation of hexose phosphates, the concentration ratio of D-glucose 6-phosphate to D-fructose 6-phosphate is much higher with the tritiated than unlabelled esters, a paradoxical increase in the specific radioactivity of D-glucose 6-phosphate above its initial value being even observed during the initial period of exposure of D-[2-3H]glucose 6-phosphate to phosphoglucoisomerase. The extension of this model to an open system may be essential for the correct interpretation of radioactive data collected in intact cells exposed to D-[2-3H]glucose.     

Murine hexose-6-phosphate dehydrogenase: a bifunctional enzyme with broad substrate specificity and 6-phosphogluconolactonase activity

Murine hexose-6-phosphate dehydrogenase has been purified from liver microsomes by affinity chromatography on 2('),5(')-ADP-Sepharose. The purified enzyme has 6-phosphogluconolactonase activity and glucose-6-phosphate dehydrogenase activity and has a native molecular mass of 178 kDa and a subunit molecular mass of 89 kDa. Glucose 6-phosphate, galactose 6-phosphate, 2-deoxyglucose 6-phosphate, glucosamine 6-phosphate, and glucose 6-sulfate are substrates for murine hexose-6-phosphate dehydrogenase, with either NADP or deamino-NADP as coenzyme. This study confirms that hexose-6-phosphate dehydrogenase is a bifunctional enzyme which can catalyze the first two reactions of the pentose phosphate pathway.     

Quantification of hepatic carbohydrate metabolism in conscious mice using serial blood and urine spots

In vivo studies of hepatic carbohydrate metabolism in (genetically modified) conscious mice are hampered by limitations of blood and urine sample sizes. We developed and validated methods to quantify stable isotope dilution and incorporation in small blood and urine samples spotted onto filter paper. Blood glucose and urinary paracetamol-glucuronic acid were extracted from filter paper spots reproducibly and with high yield. Fractional isotopomer distributions of glucose and paracetamol-glucuronic acid when extracted from filter paper spots were almost identical to those isolated from the original body fluids. Rates of infusion of labeled compounds could be adjusted without perturbing hepatic glucose metabolism. This approach was used in mice to find the optimal metabolic condition for the study of hepatic carbohydrate metabolism. In fed mice, no isotopic steady state was observed during a 6-h label-infusion experiment. In 9-h-fasted mice, isotopic steady state was reached after 3 h of label infusion and important parameters in hepatic glucose metabolism could be calculated. The rate of de novo glucose-6-phosphate synthesis was 143 +/- 17 micromol kg(-1) min(-1) and partitioning to plasma glucose was 79.0 +/- 5.2%. In 24-h-fasted mice, abrupt changes were noticed in whole body and in hepatic glucose metabolism at the end of the experiment.     

Structure-based functional annotation: yeast ymr099c codes for a D-hexose-6-phosphate mutarotase

Despite the generation of a large amount of sequence information over the last decade, more than 40% of well characterized enzymatic functions still lack associated protein sequences. Assigning protein sequences to documented biochemical functions is an interesting challenge. We illustrate here that structural genomics may be a reasonable approach in addressing these questions. We present the crystal structure of the Saccharomyces cerevisiae YMR099cp, a protein of unknown function. YMR099cp adopts the same fold as galactose mutarotase and shares the same catalytic machinery necessary for the interconversion of the alpha and beta anomers of galactose. The structure revealed the presence in the active site of a sulfate ion attached by an arginine clamp made by the side chain from two strictly conserved arginine residues. This sulfate is ideally positioned to mimic the phosphate group of hexose 6-phosphate. We have subsequently successfully demonstrated that YMR099cp is a hexose-6-phosphate mutarotase with broad substrate specificity. We solved high resolution structures of some substrate enzyme complexes, further confirming our functional hypothesis. The metabolic role of a hexose-6-phosphate mutarotase is discussed. This work illustrates that structural information has been crucial to assign YMR099cp to the orphan EC activity: hexose-phosphate mutarotase.     

The crystal structure of glucose-6-phosphate isomerase from Leishmania mexicana reveals novel active site features.

Glucose-6-phosphate isomerase catalyzes the reversible aldose-ketose isomerization of D-glucose-6-phosphate to D-fructose-6-phosphate in glycolysis and gluconeogenesis, and in the recycling of hexose-6-phosphate in the pentose phosphate pathway. The unicellular protozoans, Trypanosoma brucei, T. cruzi and Leishmania spp., of the order Kinetoplastida are important human parasites responsible for African sleeping sickness, Chagas' disease and leishmaniases, respectively. In these parasites, glycolysis is an important (and in some cases the only) metabolic pathway for ATP supply. The first seven of the 10 enzymes that participate in glycolysis, as well as an important fraction of the enzymes of the pentose phosphate pathway, are compartmentalized in peroxisome-like organelles called glycosomes. The dependence of the parasites on glycolysis, the importance of the pentose phosphate pathway in defense against oxidative stress, and the unique compartmentalization of these pathways, point to the enzymes contained in the glycosome as potential targets for drug design. The present report describes the first crystallographic structure of a parasite (Leishmania mexicana) glucose-6-phosphate isomerase. A comparison of the atomic structure of L. mexicana, human and other mammalian PGIs, which highlights unique features of the parasite's enzyme, is presented.     

Cooperativity between 11beta-hydroxysteroid dehydrogenase type 1 and hexose-6-phosphate dehydrogenase in the lumen of the endoplasmic reticulum

The functional coupling of 11beta-hydroxysteroid dehydrogenase type 1 and hexose-6-phosphate dehydrogenase was investigated in rat liver microsomal vesicles. The activity of both enzymes was latent in intact vesicles, indicating the intraluminal localization of their active sites. Glucose-6-phosphate, a substrate for hexose-6-phosphate dehydrogenase, stimulated the cortisone reductase activity of 11beta-hydroxysteroid dehydrogenase type 1. Inhibition of glucose-6-phosphate uptake by S3483, a specific inhibitor of the microsomal glucose-6-phosphate transporter, decreased this effect. Similarly, cortisone increased the intravesicular accumulation of radioactivity upon the addition of radiolabeled glucose-6-phosphate, indicating the stimulation of hexose-6-phosphate dehydrogenase activity. A correlation was shown between glucose-6-phosphate-dependent cortisone reduction and cortisone-dependent glucose-6-phosphate oxidation. The results demonstrate a close cooperation of the enzymes based on co-localization and the mutual generation of cofactors for each other.     

Effetto Del Dinitrofenolo Sulla Conversione Del Glicerolo A Glucidi in Fettine Di Carote E in Internodi Di Pisello

Abstract

On the effect of dinitrophenol on carbohydrate activation in higher plant tissues. — Previous investigations on the effects of 2,4 dinitrophenol (DNP) on carbohydrate metabolism in isolated pea internodes and in yeast showed that the increased rate of glycolysis induced by the uncoupler corresponds to an increased rate of the conversion of free hexoses and polysaccarides to hexose phosphates. In yeast about 30% of the radioactivity supplied and taken up as ¹⁴C labelled glucose, and 20% of that supplied and taken up as glycerol is recovered as soluble sugar and glycogen; this phenomenon is almost completely suppressed by 10^-4M DNP.

This suggested that a mechanism involving kinase enzymes, on one hand, and phosphatases, on the other, is mediating the interconversion of phosphorylathed and free sugars, and that the apparent increase of hexose phosphorylation observed in the presence of DNP might depend on a decreased rate of phosphatase mediate reactions, consequent to the decrease of phosphorylated sugars level in the cell.

The experiments here reported were planned to test the validity of this hypothesis in the case of higher plant tissues.

Material used in these experiments were segments from the growing part of the third internode isolated from 7 day old, etiolated pea seedlings, and carrot root diks (0,7 mm thick, 7 mm diameter) preincubated for 24 hours in aerated distilled water. Both of these materials show an active, steady respiration and some growth activity, so that they may be taken as representing a condition close enough to that of the generally physiologically active higher plant tissues.

The reversibility of the hexose phosphate-free sugar interconversion process was tested by feeding 10^-3M 1-C¹⁴ labeled glycerol, and measuring after 150 minutes the amount of radioactivity incorporated into CO₂, soluble sugars, organic acids and proteins. The results of these experiments are summarized in table I and II.

Glycerol metabolism as well as its response to DNP appears very similar in the two material used. In both cases, glycerol uptake and incorporation into organic acids and amino acids is almost insensitive to DNP. In contrast large differences are observed for the free sugar fraction. In the absence of the uncoupler, a consistent amount of the radioactivity fed as glycerol is found in this fraction. It appears reasonable to assume that the glycerol-sugar interconversion comprehends, as intermediate steps, glycerol-P, fructose di-P (or sedoeptulose di-P) and hexose-6-P. If this is true, the observed data implicate that a continuous interconversion occurs, in the cell, between sugar phosphates and free sugars and vice-versa, one reaction direction involving the activity of phosphatases, and the other one that of kinases. The true rate of this interconversion process is probably much larger than indicated by the radioactivity found in free sugars: as a considerable part of the triose-P transormed into sugars must immediately re-enter the descending flux of glycolysis.

This view finds some support in the fact that DNP almost completely inhibits the incorporation of radioactivity in the free sugar fraction. It has been previously observed that DNP very markedly decreases the level of hexose mono- and di-phosphates and of triose-phosphates in the pea stem tissues. If phosphatases acting on fructose di-phosphate and on hexose-6-P are not saturated by their substrates, a decrease of the rate of free hexose synthesis from sugar phosphates should be expected.

The present results are thus consistent with the hypothesis that hexose phosphates and free sugars in the cell are continuously interconverted by the simultaneous action of phosphatases and kinases; and that the effect of DNP, and thus of any physiological conditions decreasing the ATP/ADP ratio in accelerating free hexose utilizations is at least in part due to a decreased rate of the reactions catalized by fructose diphosphate and hexose-6-P phosphatases. The reversibility of the kinase-phosphatase system would thus represent a crucial link in the mechanism by which the rate of carbohydrate activation and breackdown is controlled by the rate of utilization of high-energy phosphate bonds.     

New reaction sequences for the non-oxidative pentose phosphate pathway.

1. Reactions leading to the formation of 14C-labelled volatile compounds and compounds volatile under acid conditions were investigated in a system actively synthesizing hexose 6-phosphates from [U-14C]ribose 5-phosphate by reactions catalysed by enzymes prepared from acetone-dried powder of rat liver; no reactions involving 14C-labelled volatile compounds were detected. Similarly the fixation of 14C-labelled volatile compounds into hexose 6-phosphate could not be detected. 2. A complete carbon balance was made for the reactants, intermediates and products of the reactions involved in the conversion of ribose 5-phosphate into hexose 6-phosphate by enzymes of rat liver. Five additional intermediates of pentose 5-phosphate metabolism in liver were detected, namely D-manno-heptulose 7-phosphate, D-altro-heptulose 1,7-bisphosphate, D-glycero-D-ido-octulose 1,8-bisphosphate, D-glycero-D-altro-octulose 1,8-bisphosphate and D-arabinose 5-phosphate. 3. D-Arabinose 5-phosphate was found to be utilized by a rat liver enzyme preparation to produce both hexose 6-phosphate and triose phosphate. 4. D-Arabinose 5-phosphate was reversibly converted into other pentose 5-phosphates. Paper chromatographic and enzymic evidence indicated that the conversion involved an enzyme tentatively named arabinose phosphate 2-epimerase, which catalyses the following reaction: D-arabinose 5-P in equilibrium D-ribose-5-P. 5. A variety of rat tissues also utilized D-arabinose 5-phosphate to produce both hexose 6-phosphate and triose phosphate and at a rate comparable with that obtained with D-ribose 5-phosphate. 6. A new reaction sequence for the non-oxidative pentose phosphate pathway in liver is proposed.     

Regulation of hysterectomy induced derangements in ovarian carbohydrate metabolism in albino rats. II. Role of prostaglandin F2 alpha.

Wistar strain albino rats were subjected to bilateral hysterectomy surgically and the ovarian carbohydrate metabolism of these animals was compared with sham operated controls. The ovarian glycogen content of hysterectomized animals was elevated with inhibition of glycogenolysis, hexose mono and diphosphate pathways and oxidative metabolism. Administration of PGF2 alpha to hysterectomized animals led to activation of ovarian glycogenolysis and other pathways of carbohydrate metabolism of hysterectomized animals was restored towards normal level after PGF2 alpha substitution.