Proportional activities of glycerol kinase and glycerol 3-phosphate dehydrogenase in rat hepatomas.

The activities of glycerol 3-phosphate dehydrogenase (EC 1.1.1.8), glycerol kinase (EC 2.7.1.30), lactate dehydrogenase (EC 1.1.1.27), "malic' enzyme (L-malate-NADP+ oxidoreductase; EC 1.1.1.40) and the beta-oxoacyl-(acyl-carrier protein) reductase component of the fatty acid synthetase complex were measured in nine hepatoma lines (8 in rats, 1 in mouse) and in the livers of host animals. With the single exception of Morris hepatoma 16, which had unusually high glycerol 3-phosphate dehydrogenase activity, the activities of glycerol 3-phosphate dehydrogenase and glycerol kinase were highly correlated in normal livers and hepatomas (r = 0.97; P less than 0.01). The activities of these two enzymes were not strongly correlated with the activities of any of the other three enzymes. The primary function of hepatic glycerol 3-phosphate dehydrogenase appears to be in gluconeogenesis from glycerol.     

Heteromerous interactions among glycolytic enzymes and of glycolytic enzymes with F-actin: effects of poly(ethylene glycol)

Interactions of glucose-6-phosphate isomerase (D-glucose-6-phosphate ketol-isomerase, EC 5.3.1.9), aldolase (D-fructose-1,6-bisphosphate D-glyceraldehyde-3-phosphate lyase, EC 4.1.2.13), glyceraldehyde-3-phosphate dehydrogenase (D-glyceraldehyde-3-phosphate:NAD+ oxidoreductase (phosphorylating), EC 1.2.1.12), triose-phosphate isomerase (D-glyceraldehyde-3-phosphate ketol-isomerase, EC 5.3.1.1), phosphoglycerate mutase (D-phosphoglycerate 2,3-phosphomutase, EC 5.4.2.1), phosphoglycerate kinase (ATP:3-phospho-D-glycerate 1-phosphotransferase, EC 2.7.3), enolase (2-phospho-D-glycerate hydro-lyase, EC 4.2.1.11), pyruvate kinase (ATP:Pyruvate O2-phosphotransferase, EC 2.7.1.40) and lactate dehydrogenase [S)-lactate:NAD+ oxidoreductase, EC 1.1.1.27) with F-actin, among the glycolytic enzymes listed above, and with phosphofructokinase (ATP:D-fructose-6-phosphate 1-phosphotransferase, EC 2.7.1.11) were studied in the presence of poly(ethylene glycol). Both purified rabbit muscle enzymes and rabbit muscle myogen, a high-speed supernatant fraction containing the glycolytic enzymes, were used to study enzyme-F-actin interactions. Following ultracentrifugation, F-actin and poly(ethylene glycol) tended to increase and KCl to decrease the pelleting of enzymes. In general, the greater part of the pelleting occurred in the presence of both F-actin and poly(ethylene glycol) and the absence of KCl. Enzymes that pelleted more in myogen preparations than as individual purified enzymes in the presence of poly(ethylene glycol) and the absence of F-actin were tested for specific enzyme-enzyme associations, several of which were observed. Such interactions support the view that the internal cell structure is composed of proteins that interact with one another to form the microtrabecular lattice.     

Enzymatic and Ultrastructural Changes in Thoracic Muscles of Triatomine Insects during the Last Stages of Metamorphosis

Dipetalogaster maximus and Triatoma infestans are hematophagous insects, vectors of Chagas' disease. After the last molt of their metamorphosis, from fifth instar nymph to adult, they acquire wings and the ability to fly, which is important for their dispersal. Some biochemical changes accompanying this last stage have been studied by determining activity of hexokinase (EC 2.7.1.1), fructose-6-phosphate kinase (EC 2.7.1.11), glucose-6-phosphate dehydrogenase (EC 1.1.1.49), glutamate dehydrogenase (EC 1.4.1.4), aspartate aminotransferase (EC 2.6.1.1), malate dehydrogenase (EC 1.1.1.37) and glycerol-3-phosphate dehydrogenase (EC 1.1.1.8) in thoracic muscle extracts of fifth instar nymphs and adults. Activity of all the enzymes, expressed in U per mg protein, was significantly higher in muscles of adults than of nymphs, except that of aspartate aminotransferase, had lower activity in adults of T. infestans. The increase of glycerol-3-phosphate dehydrogenase activity was particularly striking (30-fold), while the increase in glucose-6-phosphate dehydrogenase activity was of a lesser magnitude than those observed for other enzymes. Comparative ultrastructural studies of thoracic muscles showed that in adult preparations mitochondria were more numerous and larger in size, and presented more cristae than in muscles of fifth instar nymphs. The biochemical changes detected appear to be the expression of the adaptation of adult muscles for flight activity. Thus, adult muscles would have higher glycolytic and respiratory capacity than those of fifth instar nymphs. The operation of systems transferring hydrogen into mitochondria, especially that of the glycerophosphate shuttle, may be greatly increased in adult muscles.     

The enzymes of the classical pentose phosphate pathway display differential activities in procyclic and bloodstream forms of Trypanosoma brucei

The specific activities of each of the enzymes of the classical pentose phosphate pathway have been determined in both cultured procyclic and bloodstream forms of Trypanosoma brucei. Both forms contained glucose-6-phosphate dehydrogenase (EC 1.1.1.49), 6-phosphogluconolactonase (EC 3.1.1.31), 6-phosphogluconate dehydrogenase (EC 1.1.1.44), ribose-5-phosphate isomerase (EC 5.3.1.6) and transaldolase (EC 2.2.1.2). However, ribulose-5-phosphate 3'-epimerase (EC 5.1.3.1) and transketolase (EC 2.2.1.1) activities were detectable only in procyclic forms. These results clearly demonstrate that both forms of T. brucei can metabolize glucose via the oxidative segment of the classical pentose phosphate pathway in order to produce D-ribose-5-phosphate for the synthesis of nucleic acids and reduced NADP for other synthetic reactions. However, only procyclic forms are capable of using the non-oxidative segment of the classical pentose phosphate pathway to cycle carbon between pentose and hexose phosphates in order to produce D-glyceraldehyde 3-phosphate as a net product of the pathway. Both forms lack the key gluconeogenic enzyme, fructose-bisphosphatase (EC 3.1.3.11). Consequently, neither form should be able to engage in gluconeogenesis nor should procyclic forms be able to return any of the glyceraldehyde 3-phosphate produced in the pentose phosphate pathway to glucose 6-phosphate. This last specific metabolic arrangement and the restriction of all but the terminal steps of glycolysis to the glycosome may be the observations required to explain the presence of distinct cytosolic and glycosomal isoenzymes of glyceraldehyde-3-phosphate dehydrogenase and phosphoglycerate kinase. These same observations also may provide the basis for explaining the presence of cytosolic hexokinase and phosphoglucose isomerase without the presence of any cytosolic phosphofructokinase activity. The key enzymes of the Entner-Doudoroff pathway, 6-phosphogluconate dehydratase (EC 4.2.1.12) and 2-keto-3-deoxy-6-phosphogluconate aldolase (EC 4.1.2.14) were not detected in either procyclic or bloodstream forms of T. brucei.     

Evidence for absence of an interaction between purified 3-phosphoglycerate kinase and glyceraldehyde-3-phosphate dehydrogenase

The possibility of a functional complex formation between glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.12) and 3-phosphoglycerate kinase (EC. 2.7.2.3), enzymes catalysing two consecutive reactions in glycolysis has been investigated. Kinetic analysis of the coupled enzymatic reaction did not reveal any kinetic sign of the assumed interaction up to 4 X 10(-6) M kinase and 10(-4) M dehydrogenase. Fluorescence anisotrophy of 10(-7) M or 2 X 10(-5) M glyceraldehyde-3-phosphate dehydrogenase labeled with fluorescein isothiocynate did not change in the presence of non-labeled 3-phosphoglycerate kinase (up to 4 X 10(-5) M). The frontal gel chromatographic analysis of a mixture of the two enzymes (10(-4) M dehydrogenase) could not reveal any molecular species with the kinase activity having a molecular weight higher than that of 3-phosphoglycerate kinase. Both types of physicochemical measurements were also performed in the presence of substrates of the kinase and gave the same results. The data seem to invalidate the hypothesis that there is a complex between purified pig muscle glyceraldehyde-3-phosphate dehydrogenase and 3-phosphoglycerate kinase.     

Activity of enzymes of carbon metabolism during the induction of Crassulacean acid metabolism in Mesembryanthemum crystallinum L.

The maximum extractable activities of twenty-one photosynthetic and glycolytic enzymes were measured in mature leaves of Mesembryanthemum crystallinum plants, grown under a 12 h light 12 h dark photoperiod, exhibiting photosynthetic characteristics of either a C₃ or a Crassulacean acid metabolism (CAM) plant. Following the change from C₃ photosynthesis to CAM in response to an increase in the salinity of in the rooting medium from 100 mM to 400 mM NaCl, the activity of phosphoenolpyruvate (PEP) carboxylase (EC 4.1.1.31) increased about 45-fold and the activities of NADP malic enzyme (EC 1.1.1.40) and NAD malic enzyme (EC 1.1.1.38) increased about 4- to 10-fold. Pyruvate, Pi dikinase (EC 2.7.9.1) was not detected in the non-CAM tissue but was present in the CAM tissue; PEP carboxykinase (EC 4.1.1.32) was detected in neither tissue. The induction of CAM was also accompanied by large increases in the activities of the glycolytic enzymes enolase (EC 4.2.1.11), phosphoglyceromutase (EC 2.7.5.3), phosphoglycerate kinase (EC 2.7.2.3), NAD glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.12), and glucosephosphate isomerase (EC 2.6.1.2). There were 1.5- to 2-fold increases in the activities of NAD malate dehydrogenase (EC 1.1.1.37), alanine and aspartate aminotransferases (EC 2.6.1.2 and 2.6.1.1 respectively) and NADP glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.13). The activities of ribulose-1,5-bisphosphate (RuBP) carboxylase (EC 4.1.1.39), fructose-1,6-bisphosphatase (EC 3.1.3.11), phosphofructokinase (EC 2.7.1.11), hexokinase (EC 2.7.1.2) and glucose-6-phosphate dehydrogenase (EC 1.1.1.49) remained relatively constant. NADP malate dehydrogenase (EC 1.1.1.82) activity exhibited two pH optima in the non-CAM tissue, one at pH 6.0 and a second at pH 8.0. The activity at pH 8.0 increased as CAM was induced. With the exceptions of hexokinase and glucose-6-phosphate dehydrogenase, the activities of all enzymes examined in extracts from M. crystallinum exhibiting CAM were equal to, or greater than, those required to sustain the maximum rates of carbon flow during acidification and deacidification observed in vivo. There was no day-night variation in the maximum extractable activities of phosphoenolpyruvate carboxylase, NADP malic enzyme, NAD malic enzyme, fructose-1,6-bisphosphatase and NADP malate dehydrogenase in leaves of M. crystallinum undergoing CAM.Abbreviations CAM Crassulacean acid metabolism - PEP phosphoenolpyruvate - RuBP ribulose-1,5-bisphosphate     

Leishmania mexicana: subcellular distribution of enzymes in amastigotes and promastigotes

Glycosomes and mitochondrial vesicles from cultured promastigotes of Leishmania mexicana mexicana have been separated using isopycnic centrifugation on linear sucrose gradients. Hexokinase (EC 2.7.1.2), glucose phosphate isomerase (EC 5.3.1.9), phosphofructokinase (EC 2.7.1.11), glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.12), and phosphoenolpyruvate carboxykinase (EC 4.1.1.49) were recovered largely in association with glycosomes (density; 1.215 g/ml). Phosphoglycerate kinase (EC 2.7.2.3) and glucose-6-phosphate dehydrogenase (EC 1.1.1.49) had some small glycosomal activity, but were mostly recovered in the soluble fractions. Malate dehydrogenase (EC 1.1.1.37) showed a broad peak corresponding to that of the mitochondrial marker oligomycin-sensitive ATPase (EC 3.6.1.4) (density; 1.190 g/ml). Glutamate dehydrogenase (EC 1.4.1.3) and alanine aminotransferase (EC 2.6.1.2) both showed small mitochondrial peaks, but most of the activities were recovered elsewhere on the gradient and in the soluble fractions. The subcellular location of enzymes in L.m. mexicana amastigotes was investigated by following the release of soluble enzymes from digitonin-treated amastigotes. This revealed distinct cytosolic, mitochondrial, and glycosomal compartments. The findings give an insight into the organization and control of L.m. mexicana promastigote and amastigote energy metabolism.     

Glycerol catabolism in wild-type and mutant strains ofPseudomonas aeruginosa

Glycerol uptake, glycerol kinase (EC 2.7.1.30) and glycerol-3-phosphate dehydrogenase (EC 1.1.99.5) activities are specifically induced during growth ofPseudomonas aeruginosa PAO on either glycerol or glycerol-3-phosphate. Mutants of strain PAO unable to grow on both glycerol and glycerol-3-phosphate were isolated. Mutant PFB 121 was deficient in an inducible, membrane-bound, pyridine nucleotide-independent, glycerol-3-phosphate dehydrogenase activity and PFB 82 was deficient in glycerol uptake and glycerol kinase and glycerol-3-phosphate dehydrogenase activities. Each mutant spontaneously reverted to wild phenotype, which indicates that each contained a single genetic lesion. These results demonstrate that membrane-bound, inducible glycerol-3-phosphate dehydrogenase is required for catabolism of both glycerol and glycerol-3-phosphate and provide suggestive evidence for a single regulatory locus that controls the synthesis of glycerol uptake, glycerol kinase, and glycerol-3-phosphate dehydrogenase inP. aeruginosa.     

Triiodothyronine depresses the NAD-linked glycerol-3-phosphate dehydrogenase activity of cultured neonatal rat heart cells

The activity of NAD-linked alpha-glycerol-3-phosphate dehydrogenase (NAD-G3PDH; EC 1.1.1.8) was depressed by 35% when the thyroid hormone 3,3',5-triiodo-L-thyronine (20 micrograms/liter) was added to the serum-free, hormonally supplemented medium of cultured neonatal rat heart cells. The degree of depression was greater (65%) when the medium contained normal serum levels of hydrocortisone and insulin. There is a dramatic inverse dose-response relationship between triiodothyronine levels and NAD-G3PDH activity. The classic elevation by thyroid hormones of the FAD-linked alpha-glycerol-3-phosphate dehydrogenase (FAD-G3PD; EC 1.1.99.5) was observed concurrently. The medium-glucose depletion rate in triiodothyronine-free cells was depressed 32% through 11 days-in-culture, indicating reduced glycolytic activity. The activities of nine other metabolically important enzymes which were measured during this study, including hexokinase, glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, phosphofructokinase, pyruvate kinase, malate dehydrogenase, NAD-isocitrate dehydrogenase, NADH cytochrome c reductase, and succinic cytochrome c reductase, did not respond to varying triiodothyronine concentrations.     

Plagiorchis elegans: Histochemical localization of dehydrogenases in the cercarial stage

Cercariae of Plagiorchis elegans Rudolphi 1802 collected from experimentally infected snails, Lymnaea palustris, were subjected to various histochemical tests for dehydrogenase systems. A high degree of activity was demonstrated for succinic dehydrogenase (EC 1.3.99.1), malic dehydrogenase (EC 1.1.1.37), isocitric dehydrogenase (EC 1.1.1.41), α-glycerophosphate dehydrogenase (EC 1.1.1.8), and glucose 6-phosphate dehydrogenase (EC 1.1.1.49). These enzymes were present in the tegument, tail, caudal pocket, excretory bladder, acetabulum, and oral sucker, particularly in the muscles around the stylet. Only moderate activity was obtained for lactic dehydrogenase (EC 1.1.1.27) and 6-phosphogluconate dehydrogenase (EC 1.1.1.44) at these sites, glutamic dehydrogenase (EC 1.4.1.2) was localized only in the tails of the cercariae and tests for alcohol dehydrogenase (EC 1.1.1.1) were completely negative. The cerebral ganglia and its commissures stained intensely in the tests for succinic, isocitric, α-glycerophosphate, and glucose 6-phosphate dehydrogenase systems. The results indicate the possibility that several energy-producing sequences may be available to these cercariae.     

Separation of Neurospora Crassa Myo-Inositol-1-Phosphate Synthase from Glucose-6-Phosphate Dehydrogenase by Affinity Chromatography

The purification of Neurospora crassa myo-inositol-1-phosphate synthase (EC 5.5.1.4) was studied by affinity chromatography using the substrate (glucose-6-phosphate), the inhibitor (pyrophosphate), the coenzyme (NAD⁺) and the coenzyme analogues (5′AMP and Cibacron Blue F3G-A) of the enzyme as adsorbents attached to agarose gel. Myo-inositol-1-phosphate synthase could be separated completely from the contaminating substance, glucose-6-phosphate dehydrogenase (EC 1.1.1.49), on Blue Sepharose CL-6B and on pyrophosphate-Sepharose. The purified enzyme had a specific activity of 16 400 U/mg. The sodium dodecyl sulfate/polyacrylamide gel electrophoresis of 60 μq of this purified enzyme gave a homogenous band. The enzyme was found to be composed of four identical subunits having a molecular weight of 65 000.     

Measurement of the inorganic pyrophosphate in tissues of Pisum sativum L.

Purified pyrophosphate: fructose 6-phosphate 1-phosphotransferase (EC 2.7.1.90) was used to measure the inorganic pyrophosphate in unfractionated extracts of tissues of Pisum sativum L. The fructose 1,6-bisphosphate produced by the above enzyme was measured by coupling to NADH oxidation via aldolase (EC 4.1.2.13), triosephosphate isomerase (EC 5.3.1.1) and glycerol-3-phosphate dehydrogenase (EC 1.1.1.8). Amounts of pyrophosphate as low as 1 nmol could be measured. The contents of pyrophosphate in the developing embryo of pea, and in the apical 2 cm of the roots, were appreciable; 9.4 and 8.9 nmol g^-1 fresh weight, respectively. The possibility that pyrophosphate acts in vivo as an energy source for pyrophosphate: fructose 6-phosphate 1-phosphotransferase and for UDPglucose pyrophosphorylase (EC 2.7.7.9) is considered.     

Enzymes of starch and sugar phosphate metabolism in achlorophyllous ribosome-deficient plastids from high-temperature-grown rye leaves

Several enzymes of non–photosynthetic sugar phosphate and starch metabolism were measured in gradient–purified chloroplasts from normal rye leaves ( Secale cereale L. cv. Halo) grown at 22°C and in the non-photosynthetic plastids isolated from 70S ribosome-deficient rye leaves grown at a non–permissive elevated temperature of 32°C. Activities of the enzymes phosphoglycerate kinase (EC 2.7.2.3), hexokinase (EC 2.7.1.1), phosphoglucose isomerase (EC 5.3.1.9), phosphoglucomutase (EC 2.7.5.1), glucose-6-phosphate dehydrogenase (EC 1.1.1.49), 6-phosphogluconate de-hydrogenase (EC 1.1.1.46), ADPglucose pyrophosphorylase (EC 2.7.7.27), starch synthase (EC 2.4.1.21), and phosphorylase (EC 2.4.1.1) were present in ribosome-deficient plastids from 32°C-grown leaves indicating a cytoplasmic origin of the plastid-specific forms of these enzymes. While the photosynthetic marker enzyme NADP⁺-dependent glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.13) was considerably diminished, both the specific activities and the total activities per leaf of the plastid-specific forms of hexokinase, phosphoglucose isomerase and phosphoglucomutase were markedly increased in the ribosome–deficient plastids, relative to normal chloroplasts. The results demonstrate that after elimination of functional protein synthesis in the chloroplasts the supply of chloroplast–specific enzymes by the cytoplasm is not generally suppressed as observed for many enzymes and proteins involved in photosynthesis, but may even be increased in accord with changed metabolic demands.     

Enzymatic synthesis of UDP-galactose on a gram scale

Uridine 5′-diphospho-- -galactose (UDP-Gal) was synthesized on a gram scale from uridine 5′-diphospho-- -glucose and - -galactose 1-phosphate using the enzymes galactose-1-phosphate uridyltransferase (EC 2.7.7.12), phosphoglucomutase (EC 2.7.5.1) and glucose-6-phosphate dehydrogenase (EC 1.1.1.27). The synthesis was performed in a repetitive batch mode in which the enzymes, some of which are expensive, were used in 16 subsequent batches without any loss of enzyme activity. The space time yield of the synthesis was 7.1 g/l d. The overall yield of the synthesis amounted to 40% and 1.1 gram of pure UDP-Gal was obtained.     

Selective denaturation of several yeast enzymes by free fatty acids.

The denaturation of eight purified yeast enzymes, glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, glyceraldehyde-3-phosphate dehydrogenase, 3-phosphoglycerate kinase, alcohol dehydrogenase, beta-fructosidase, hexokinase and glucose-6-phosphate isomerase, promoted under controlled conditions by the free fatty acids myristic and oleic, is selective. Glucose-6-phosphate dehydrogenase (D-glucose-6-phosphate:NADP+ 1 oxidoreductase, EC 1.1.1.49) is extremely sensitive to destabilization and was studied in greater detail. Results show that chain length and degree of unsaturation of fatty acids are important to their destabilizing effect, and that ligands of the enzyme can afford protection. The denaturation process results in more than one altered form. These results can be viewed in the perspective of the possibility that amphipathic substances, and in particular free fatty acids, may play a role for enzyme degradation in vivo, by initiating steps of selective denaturation.     

Polymorphism of several dehydrogenases in the skin of sheep and their fetuses

The total activity and isoenzyme spectra of lactate dehydrogenase (EC 1.1.1.27), malate dehydrogenase (EC 1.1.1.37), glutamate dehydrogenase (EC 1.4.1.2) and glucose-6-phosphate dehydrogenase (EC 1.1.1.49) were studied in the skin of sheep and their fetuses. There are 5 multiple molecular forms of LDG, 4--of GDG, 2--of MDG and 1--of G-6-PDG in the skin. It is established that the skin of adult sheep and fetus is similar as to the studied parameters.     

Schistosomatium douthitti: carbohydrate metabolism in the adults.

Pathways of carbohydrate metabolism in the adults of Schistosomatium douthitti: were investigated. Histochemical reactions for adenosinetriphosphatase (EC 3.6.1.3) glucose 6-phosphate dehydrogenase (EC 1.1.1.49), phosphogluconate dehydrogenase (EC 1.1.1.43), glycerol-3-phosphate dehydrogenase (EC 1.1.1.8), lactate dehydrogenase (EC 1.1.1.27, 1.1.2.3) isocitrate dehydrogenase (EC 1.1.1.41), succinate dehydrogenase (EC 1.3.99.1), malate dehydrogenase (EC 1.1.1.37), cytochrome oxidase (EC 1.9.3.1), and adenosine triphosphatase (EC 3.6.1.3) were found in the adult worms. Glycogen deposits occurred in the parenchyma.Low oxygen tension immobilized the worms. Tartar emetic, sodium cyanide reduced adult motility in vitro. Manometric experiments demonstrated a respiratory quotient of approximately one. Oxygen uptake was completely inhibited by tartar emetic and partially inhibited by sodium fluoracetate and sodium cyanide. Inhibition by sodium fluoroacetate was partially counteracted by citric acid in the medium.Adults demonstrated an oxygen debt following anaerobic incubation. A maximum of 52% of the glucose consumed under aerobic conditions was excreted as lactic acid. Under anaerobic conditions the amount of lactic acid excreted increased. Acids other than lactic acid were also released. Results indicate that although glycolysis is the major pathway, two additional aerobic pathways also exist, one which is cyanide sensitive and the other cyanide insensitive.     

Separation of Neurospora crassa myo-inositol-1-phosphate synthase from glucose-6-phosphate dehydrogenase by affinity chromatography

The purification of Neurospora crassa myo-inositol-1-phosphate synthase (EC 5.5.1.4) was studied by affinity chromatography using the substrate (glucose-6-phosphate), the inhibitor (pyrophosphate), the coenzyme (NAD+) and the coenzyme analogues (5'AMP and Cibacron Blue F3G-A) of the enzyme as adsorbents attached to agarose gel. Myo-inositol-1-phosphate synthase could be separated completely from the contaminating substance, glucose-6-phosphate dehydrogenase (EC 1.1.1.49), on Blue Sepharose CL-6B and on pyrophosphate-Sepharose. The purified enzyme had a specific activity of 16 400 U/mg. The sodium dodecyl sulfate/polyacrylamide gel electrophoresis of the 60 micrograms of this purified enzyme gave a homogenous band. The enzyme was found to be composed of four identical subunits having a molecular weight of 65 000.     

Mannitol production in fungi during glucose catabolism.

The levels of phosphofructokinase (EC 2.7.1.11) and mannitol-1-phosphate dehydrogenase (EC 1.1.1.17) have been determined in a number of Mucor and Penicillium species. Mannitol-1-phosphate dehydrogenase was found in only one species of mucor, Mucor rouxii, and this with a specific activity much lower than that found in Penicillium species. All of the fungi tested in the Ascomycetes class exhibited mannitol-1-phosphate dehydrogenase activity. Interference from both mannitol-1-phosphate dehydrogenase and NADH oxidase (EC 1.6.99.5) caused some difficulty initially in detecting phosphofructokinase in Penicillium species; the Penicillium phosphofructokinase is very unstable. Penicillium notatum accumulates mannitol intracellularly; detection of mannitol-1-phosphate dehydrogenase and mannitol-1-phosphatase (EC 3.1.3.22) activity in cell-free extracts indicates that the mannitol is formed from glucose via fructose-6-phosphate and mannitol-1-phosphate; no direct reduction of fructose to mannitol could be detected. The mannitol-1-phosphate dehydrogenase was specific for mannitol-1-phosphate and fructose-6-phosphate; NADP+(H) could not replace NAD+(H). The phosphatase (EC3.1.3.22) exhibited a distinct preference for mannitol-1-phosphate as substrate; all other substrates tested exhibited less than 25% of the activity observed with mannitol-1-phosphate.     

Specific activities of 6-phosphogluconate dehydrogenase,glucose-6-phosphate dehydrogenase and glucose-6-phosphate isomerase during Bufo bufo development

It has been suggested by some authors that during amphibian development, due to the higher glucose-6-phosphate dehydrogenase (EC 1.1.1.49) activity compared to that of 6-phosphogluconate dehydrogenase (EC 1.1.1.43), 6-phosphogluconate could accumulate in the embryo tissues and regulate the channelling of glucose-6-phosphate into glycolysis. Here, on the base of the specific activities of glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase and glucose-6-phosphate isomerase (EC 5.3.1.9) found in the embryos of Bufo bufo during development, it is discussed whether 6-phosphogluconate can accumulate and play a regulative role on glucose-6-phosphate metabolism in the anuran embryo.