Control analysis as a tool to understand the formation of the las operon in Lactococcus lactis

In Lactococcus lactis the enzymes phosphofructokinase (PFK), pyruvate kinase (PK) and lactate dehydrogenase (LDH) are uniquely encoded in the las operon. We used metabolic control analysis to study the role of this organization. Earlier studies have shown that, at wild-type levels, LDH has no control over glycolysis and growth rate, but high negative control over formate production (C(Jformate)LDH=-1.3). We found that PFK and PK exert no control over glycolysis and growth rate at wild-type enzyme levels but both enzymes exert strong positive control on the glycolytic flux at reduced activities. PK exerts high positive control over formate (C(Jformate)PK=0.9-1.1) and acetate production (C(Jacetate)PK=0.8-1.0), whereas PFK exerts no control over these fluxes at increased expression. Decreased expression of the entire las operon resulted in a strong decrease in the growth rate and glycolytic flux; at 53% expression of the las operon glycolytic flux was reduced to 44% and the flux control coefficient increased towards 3. Increased las expression resulted in a slight decrease in the glycolytic flux. At wild-type levels, control was close to zero on both glycolysis and the pyruvate branches. The sum of control coefficients for the three enzymes individually was comparable with the control coefficient found for the entire operon; the strong positive control exerted by PK almost cancels out the negative control exerted by LDH on formate production. Our analysis suggests that coregulation of PFK and PK provides a very efficient way to regulate glycolysis, and coregulating PK and LDH allows cells to maintain homolactic fermentation during glycolysis regulation.     

Reversible phosphorylation control of skeletal muscle pyruvate kinase and phosphofructokinase during estivation in the spadefoot toad,Scaphiopus couchii

Both pyruvate kinase (PK) and phosphofructokinase (PFK) occur in two different forms, separable by isoelectric focusing (IEF), in skeletal muscle of the spadefoot toad Scaphiopus couchii. During estivation (aerobic dormancy) the proportions of the two forms changed compared with controls; in both cases the amount of enzyme in Peak I (pI = 5.3-5.4) decreased whereas activity in Peak II (isoelectric point = 6.2-6.4) increased. In vitro incubation of crude muscle extracts with 32P-ATP under conditions that promoted the activity of cAMP-dependent protein kinase led to strong radiolabeling associated with Peak I, but not Peak II, and reverse phase HPLC confirmed that 32P was associated with the subunits of both PK and PFK found in Peak I. Specific radiolabeling of Peak I PK and PFK by protein kinase A was further confirmed using immunoprecipitation. In total, this information allowed identification of the Peaks I and II enzymes as the phosphorylated and dephosphorylated forms, respectively, and the effect of estivation was to increase the proportion of dephosphorylated PK and PFK in muscle. Analysis of the kinetic properties of partially purified PK and PFK revealed significant kinetic differences between the two forms of each enzyme. For PK, the Peak II (low phosphate) enzyme showed a 1.6-fold higher Km for phosphoenolpyruvate and a 2.4-fold higher Ka for fructose-1,6-bisphosphate than did the Peak I (high phosphate) form. These kinetic properties suggest that Peak II PK is the less active form, and coupled with the shift to predominantly the Peak II form during estivation (87% Peak II vs. 13% Peak I), are consistent with a suppression of PK activity in estivating muscle, as part of the overall metabolic rate depression of the estivating state. A similar shift to predominantly the Peak II, low phosphate, form of PFK (75% Peak II, 25% Peak I) in muscle of estivating animals is also consistent with metabolic suppression since phosphorylation of vertebrate skeletal muscle PFK is typically stimulated during exercise to enhance enzyme binding to myofibrils in active muscle. Peak II PFK also showed reduced sensitivity to inhibition by Mg:ATP (I50 50% higher) compared with the Peak I form suggesting that the enzyme in estivating muscle is less tightly regulated by cellular adenylate status than in awake toads. The data indicate that reversible phosphorylation control over the activity states of enzymes of intermediary metabolism is an important mechanism for regulating transitions between dormant and active states in estivating species.     

Lactate downregulates the glycolytic enzymes hexokinase and phosphofructokinase in diverse tissues from mice

We examined the effects of lactate on the enzymatic activity of hexokinase (HK), phosphofructokinase (PFK) and pyruvate kinase (PK) in various mouse tissues. Our results showed that lactate inhibited PFK activity in all the analyzed tissues. This inhibitory effect was observed in skeletal muscle even in the presence of insulin. Lactate directly inhibited the phosphorylation of PFK tyrosine residues in skeletal muscle, an important mechanism of the enzyme activation. Moreover, lactate indirectly inhibited HK activity, which resulted from its cellular redistribution, here attributed to alterations of HK structure. PK activity was not affected by lactate. The activity of HK and PFK is directly related to glucose metabolism. Thus, it is conceivable that lactate exposure can induce inhibition of glucose consumption in tissues.     

Characterization of Key Glycolytic and Oxidative Enzymes in Steinernema carpocapsae

The enzyme activities of isocitrate dehydrogenase (ICDH, NADP-specific), lactate dehydrogenase (LDH), malate dehydrogenase (MDH), phosphoenolpyruvate carboxykinase (PEPCK), phosphofructokinase (PFK), pyruvate kinase (PK), and fructose-l,6-bisphosphatase (FBPase) were studied in the third-stage juveniles of Steinernema carpocapsae. Reaction requirements, pH optima, substrate and cofactor kinetic constants were similar to those reported previously from other parasitic helminths with the exception of LDH, which was unstable and could not be characterized for specific activity and kinetic constants. The respective pH optima were 7.5 for ICDH, 8.8 for MDH, 6.5 for PEPCK, 7.3 for PFK, 7.2 for PK, and 7.5 for FBPase. The specific activities for ICDH, MDH, PEPCK, PFK, PK, and FBPase at pH 7.5 were 4.8, 1,300, 22, 25, 35, and 6.8 (nmoles substrate ∙ min⁻¹ ∙ mg protein⁻¹), respectively. In summary, the infective juveniles of S. carpocapsae display the metabolism typical of a facultative aerobe.     

Temperature alterations during embryogenesis have a sex-dependent influence on growth properties and muscle metabolism of day-old chicks and 35-day-old broilers

Broiler eggs were either incubated at 37.8°C during the whole incubation period (control), or at higher (38.8°C, group H) and lower temperatures (36.8°C, group L) from embryonic day (ED) 7 up to ED 10 (ED 7 to 10) or from ED 10 up to ED 13 (ED 10 to 13). Before and after this temperature treatment the eggs were incubated at 37.8°C. The day-old chicks were weighted, sexed and fed up to day 35. On days 1 and 35 samples were taken from the breast and leg muscles for analyzing of the mitochondrial respiratory activity (MRA) and from the breast muscles for analysis of the cross-sectional areas (CSA) and the glycogen phosphorylase (GP), phosphofructokinase (PFK), lactate dehydrogenase (LDH), citrate synthase (CS) and cytochrome oxidase (COX) activities. Statistical analysis showed that treatment (control, group H, group L), sex and their interaction, but not the treatment period (ED 7 to 10; ED 10 to 13), significantly influenced the results. Group H chicks had lower (P⩽0.05) body and heart weights but higher (P⩽0.05) liver weights, CSA values, leg MRA as well as PFK, LDH, CS, GP and COX activities compared with the group L chicks. The results of the control chicks differ (P⩽0.05) from those of the group H (body, heart weight, COX), the group L chicks (liver weight, PFK, LDH, CS, GP) or the birds of both other groups (CSA). The group H broiler had higher (P⩽0.05) body and leg weights as well as LDH, CS, COX and GP activities than the group L broilers. The BWs and the LDH and GP results of the control broiler differ (P⩽0.05) from those of both other groups or from the results of the group H (CS) and group L broiler (COX). Female broilers had lower (P⩽0.05) body, breast and leg weights, but higher (P⩽0.05) CSA, LDH, CS and GP activities than the male animals. Analysis of treatment×sex interaction showed that group H hens had higher (P⩽0.05) body and breast weights, LDH and GP activities compared with the group L hens, whereas in the male broiler no effect of the interaction could be found, except for the lower (P⩽0.05) CSA values in the group H than group L cocks. The treatment effects are probably due to altered embryonic activity and related molecular mechanisms. The sex-related differences in the broiler indicate that these alterations already occur in the embryos and chicks, but become significant with the sexual dimorphism after hatch.     

Paralysis of Innervated Cultured Human Muscle Fibers Affects Enzymes Differentially

Increased accumulation of muscle-specific isozyme (MSI) of creatine kinase (CK), lactate dehydrogenase (LDH), glycogen phosphorylase (GP), and phosphoglycerate mutase (PGAM) occurs with development and indicates muscle fiber maturation. The expression of MSIs of those four enzymes is greatly enhanced in innervated-contracting as compared to noninnervated and noncontracting cultured human muscle fibers. We have now studied the effect of contractile activity on developmental accumulation of MSIs in innervated-contracting, innervated-paralyzed (2 microM tetrodotoxin for 30 days), and noninnervated-noncontracting cultured human muscle fibers. Muscle acetylcholinesterase (AChE) and total enzyme activities were also studied under the same conditions. We observed a different dependency on contractile activity between total enzymatic activities of CK, LDH, and AChE, which were substantially reduced after paralysis, and GP and PGAM, which were unchanged. The expression of MSIs of CK, GP, PGAM, and LDH was always significantly increased in innervated as compared to noninnervated fibers. While the expression of MSIs of GP and PGAM was the same in contracting-innervated and paralyzed-innervated muscle fibers, the expression of MSIs of CK and LDH in paralyzed-innervated muscle fibers was very slightly decreased as compared to their contracting-innervated controls. Our studies demonstrate that in human muscle: (1) total enzymatic activities and the expression of MSIs of GP and PGAM are regulated by neuronal effect(s); (2) total enzymatic activities of CK, LDH, and AChE depend mainly on muscle contractile activity; and (3) MSIs of CK and LDH are regulated predominantly by neuronal factors and to a much lesser degree by muscle contractile activity.     

Development of the activities of oxidative, glycolytic and muscle enzymes during early larval life in three families of freshwater fish

The development of the activities of oxidative (COX, CS), glycolytic (PFK, PK, LDH) and muscle enzymes (CK, MK, Pase) was studied in representatives of the families Coregonidae, Salmonidae and Cyprinidae, from hatching to an age of approximately 100 days. In addition, the activities of two enzymes of amino acid metabolism (GOT, GPT) were followed in rainbow trout and in roach.
Water content of fresh body weight and protein content of dry body weight decrease during the early larval period. Specific activities of the two oxidative enzymes decline, whereas those of glycolytic and muscle enzymes increase in all species.
A family-specific event is the enormous increase in glycolytic and muscle enzymes from very low values in the early larva to very high levels in adult Coregonus sp. In rainbow trout, CS activity begins with a low-level period lasting throughout the yolk-sac period, whereas in the other species CS activity is high immediately after hatching.
Acclimation to either 15 or 20° C has no effect on the mass-specific activities of PFK, M K, CK and Pase in roach and chub, but the former three enzymes appear to be strongly dependent on rearing conditions during the early larval period, whereas Pase is not.     

Effect of copper on liver key enzymes of anaerobic glucose metabolism from freshwater tropical fish Prochilodus lineatus

We investigated the effect of copper on liver key enzymes of the anaerobic glucose metabolism (hexokinase, HK; phosphofructokinase, PFK; pyruvate kinase, PK; lactate dehydrogenase, LDH) as well as of the pentose pathway (glycose-6-phosphate dehydrogenase, G6PDH) from the fish Prochilodus lineatus. The fish were acclimated at either 20 degrees C or 30 degrees C at pH 7.0, transferred to water at pH 4.5 or 8.0, and exposed to 96 h-CL(50) copper concentrations. Copper accumulation in liver was higher in fish acclimated at 20 degrees C and maintained in water pH 8.0. Three-way analysis of variance revealed a significant effect of temperature on all enzymes, a significant effect of pH on all enzymes except for PK, and a significant effect of copper on only PFK, and LDH in pH 4.5 at 20 degrees C and, at 30 degrees C, on PFK and PK at pH 4.5 and 8.0, HK at pH 4.5 and G6PDH at pH 8.0. There were significant interactions between treatments for many enzymes. These changes suggest that the activity of enzymes in question is modified by a change in ambient water. At least at 30 degrees C, the overall reduction in the glycolytic enzyme activities of copper-exposed fish seems to reduce energy availability via glucose metabolism, thereby contributing to enhance copper toxic effects.     

Effect of carcinogenic and non-carcinogenic chemicals on the activities of four glycolytic enzymes in mouse lung

The activities of four glycolytic enzymes were measured in the lung homogenate of CFLP mice treated with a variety of carcinogens and non-carcinogens for mouse lung. The carcinogenic urethane, dimethylnitrosamine (DMNA), 3-methylcholanthrene (MCA), benzo[a]pyrene (BP), 7,12-dimethylbenz[a]anthracene (DMBA) and aflatoxin B1 enhanced the activity of hexokinase (HK), phosphofructokinase (PFK), pyruvate kinase (PK) and lactate dehydrogenase (LDH) 28 days after a single intraperitoneal administration. These carcinogens also altered the ratio of LDH H and M subunits. In contrast, under the same conditions the non-carcinogenic phenylurethane, ethylformate, chrysene, perylene and pyrene, as well as the pulmonarily toxic Paraquat, butylated hydroxytoluene (BHT) and cadmium chloride (CdCl2), did not influence either the activities of the enzymes tested or the isozyme pattern of LDH.     

Carbohydrate metabolism in temporal and persistent hypoglycemic chickens induced by insulin infusion

In order to elucidate the regulatory mechanism of blood glucose concentrations specific to chickens, carbohydrate metabolism in the liver, muscle and kidney and metabolite concentrations in the blood were investigated in chickens with acute and persistent hypoglycemia. Acute and persistent hypoglycemia were experimentally induced by a single injection of insulin (8 U/kg BW) or by continuous infusion of insulin (22.5 U/kg BW/day) for 4 days. Non-esterified fatty acid (NEFA) concentration in plasma and D-3-hydroxybutyrate (3HB) concentrations in liver and muscle increased in the acute hypoglycemia. Plasma NEFA concentration and 3HB concentration in the blood and liver were not changed at day 3 of persistent hypoglycemia, while 3HB concentration in the muscle was decreased. Phosphofructokinase (PFK) activity in the liver tended to increase but PFK and pyruvate kinase (PK) activities were unchanged in acute hypoglycemia. In persistent hypoglycemia, increase of hepatic PFK activity at day 1 in which it was reversed at day 3, and a small increase of muscle PK activity were observed, while PK and phosphoenolpyruvate carboxykinase (PEPCK) activities in the liver and kidney were not significantly changed. These results show that in the persistent hypoglycemic chickens, hepatic glycolysis transiently increases, which is followed by a small decrease, while glycolysis in muscles and gluconeogenesis in the liver and kidney are not significantly changed.     

The influence of muscle metabolic characteristics on physical performance

This study describes the influence of muscle fiber type composition, enzyme activities and capillary supply on muscle strength, local muscle endurance or aerobic power and capacity. Muscle biopsies were obtained from m. vastus lateralis in thirteen physically active men. Histochemical staining procedures were applied to assess the percentage of fast twitch (FT) fibers, muscle fiber area, and capillary density. Also, the activity of citrate synthase (CS), creatine kinase (CK), hexokinase (HK), lactate dehydrogenase (LDH), and phosphofructokinase (PFK) were analysed using fluorometrical assays. Peak torque at 'low' and 'high' angular velocities was measured during leg extension. Similarly, muscle fatigue (e.g. peak torque decline) and recovery from a short-term exercise task were measured during maximal, voluntary consecutive leg extensions. Aerobic power (VO2max) and aerobic capacity (e.g. onset of blood lactate concentration; OBLA), as defined by a blood lactate concentration of 4 mol X 1(-1) were measured during cycling. Peak torque at a high angular velocity was positively correlated with % FT area (p less than 0.001). Fatigue and recovery were correlated with LDH X CS-1 (p less than 0.001). WOBLA was best correlated with PFK and PFK X CS-1 (p less than 0.001). Hence, muscle strength was partly determined by fiber type composition whereas local muscle endurance, recovery and aerobic capacity reflect mainly capillary supply and the activity of key enzymes involved in aerobic and anaerobic metabolism.     

Mechanisms of glycolytic control during hibernation in the ground squirrel Spermophilus lateralis

Summary The mechanisms of glycolytic rate control during hibernation in the ground squirrel Spermophilus lateralis were investigated in four tissues: heart, liver, kidney, and leg muscle. Overall glycogen phosphorylase activity decreased significantly in liver and kidney to give 50% or 75% of the activity found in the corresponding euthermic organs, respectively. The concentration of fructose-2,6-bisphosphate (F-2,6-P₂) decreased significantly in heart and leg muscle during hibernation to 50% and 80% of euthermic tissue concentrations, respectively, but remained constant in liver and kidney. The overall activity of pyruvate dehydrogenase (PDH) in heart and kidney from hibernators was only 4% of the corresponding euthermic values. Measurements of phosphofructokinase (PFK) and pyruvate kinase (PK) kinetic parameters in euthermic and hibernating animals showed that heart and skeletal muscle had typical rabbit skeletal M-type PFK and M₁-type PK. Liver and kidney PFK were similar to the L-type enzyme from rabbit liver, whereas liver and kidney PK were similar to the M₂ isozyme found primarily in rabbit kidney. The kinetic parameters of PFK and PK from euthermic vs hibernating animals were not statistically different. These data indicate that tissue-specific phosphorylation of glycogen phosphorylase and PDH, as well as changes in the concentration of F-2,6-P₂ may be part of a general mechanism to coordinate glycolytic rate reduction in hibernating S. lateralis.Abbreviations ADP adenosine diphosphate - AMP adenosine monophosphate - ATP adenonine triphoshate - EDTA ethylenediaminetetra-acetic acid - EGTA ethylene glycol tetra-acetic acid - F-6-P fructose 6-phosphate - F-1,6-P₂ fructose 1,6-bisphosphate - F-2,6-P₂ fructose-2,6-bisphosphate - K _a activation coefficient - I₅₀ concentration of inhibitor which reduces control activity by 50% - PDH pyruvate dehydrogenase - PEP phosphoenolpyruvate - PFK 6-phosphofructo-1-kinase - PK pyruvate kinase     

Metabolic differentiation of homologous leg muscles of two aquatic birds at the level of enzymatic organization

The quantitative determination of succinic dehydrogenase (SDH), hexokinase (HK), phosphorylase, phosphofructokinase (PFK), glycerol-3-phosphate dehydrogenase (G-3-PDH) and lactate dehydrogenase (LDH) was carried out in the homologous leg muscles of two aquatic Birds. It appears that the leg muscle fibres of the coot, a surface swimmer are more oxidative in nature and appear to utilize glucose as source of energy. The leg muscles of the dabchick, a diving Bird, on the other hand, seem to depend on glycogen as source of energy. The relative activity levels of HK, phosphorylase and PFK support the accepted r?le of glycogen as primary substrate of carbohydrate catabolism in the leg muscles. The ratio of G-3-PDH/LDH in the leg muscles revealed that glycerol 3-phosphate cycle appears to be insufficient to account for the major part of NADH oxidation. However, the LDH activity is quite high in all the muscles. These results led us to believe that glycerol 3-phosphate cycle may function during rest, when the rate of glycolysis will be low.     

Effect of exercise on glycolytic enzymes of zucker fatty rats

Summary The effect of fatigue (running to exhaustion) on the Vmax activity of the key glycolytic enzymes measured at saturating substrate concentrations in muscles, liver and brain of sedentary and trained (running on a treadmill one h/day at 20 m/min, five days/week for six months) female Zucker fatty rats and their lean littermates was investigated. In the sedentary rats, fatigue increased the activity of phosphofructokinase (PFK) in the red vastus muscle by 82% in lean, and 120% in obese rats. In the trained rats, fatigue increased PFK activity by 28% in the white vastus muscle of lean rats. In the lean animals, hexokinase (HK) activity was decreased by 26% in the red vastus of sedentary rats, and by 29% in the white vastus of trained rats upon fatiguing. Pyruvate kinase (PK) activity was also decreased by 29% in the white vastus of fatigued lean animals. Training by itself had no effect on the activity of glycolytic enzymes, except PK activity which was increased by 27% in the cortex of the lean animals. It is concluded that in the Zucker rat, these glycolytic enzymes may play a differential role in regulating glycolysis during exercise and fatigue; the extent of their involvement differs depending upon the type of tissue studied and exercise. In view of the reported short half-life (7–17 h) of PFK and its covalent modification, it is suggested that the total content and/or phosphorylation status of the enzyme may be affected in animals subjected to long-term fatigue.Abbreviations PFK Phosphofructokinase (EC 2.7.1.11) - PK Pyruvate Kinase (EC 2.7.1.40) - HK Hexokinase (EC 2.7.1.1) - LSC Lean Sedentary Control - LTC Lean Trained Control - LSF Lean Sedentary Fatigued - LTF Lean Trained Fatigued - OSC Obese Sedentary Control - OTC Obese Trained Control - OSF Obese Sedentary Fatigued - OTF Obese Trained Fatigued     

Longitudinal and allometric variation in indicators of muscle metabolic capacities in atlantic cod (Gadus morrhua)

This study evaluated whether indicators of metabolic capacity of cod white muscle differ along the length of the body, whether this variation persists over a large range of body sizes, and whether the allometry of metabolic capacities is similar along the length of the body. We examined the maximal activities of two glycolytic enzymes, phosphofructokinase (PFK) and lactate dehydrogenase (LDH), a mitochondrial enzyme, cytochrome C oxidase (CCO), and the biosynthetic enzyme nucleotide diphosphate kinase (NDPK). All enzymes examined showed significant size dependence, which was generally apparent in all regions. The activity of glycolytic enzymes increased with size, whereas that of CCO and NDPK decreased with size. For PFK and LDH, the size dependence decreased caudally, whereas for CCO and NDPK it was strongest in the caudal sample. For each size range, the activities of PFK, LDH, and CCO were higher in the last third of the body than in the middle or just behind the head. In contrast, NDPK activity was higher just behind the head than at the middle or in the last third of the body, suggesting that nuclear proliferation is more rapid in this zone. The high capacity for adenosine triphosphate (ATP) generation in the caudal region suggests that increases in mass-specific ATP output are advantageous in this relatively thin section of the body.     

From genome to enzyme: analysis of key glycolytic and oxidative pentose-phosphate pathway enzymes in the cyanobacterium Synechocystis sp. PCC 6803

Activities of glucokinase, glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, phosphoglucose isomerase, phosphofructokinase (PFK), enolase, pyruvate kinase (PK) and phosphoenolpyruvate (PEP) carboxylase were determined in extracts of photoautotrophic, mixotrophic, and heterotrophic cultures of Synechocystis sp. PCC 6803. Annotated genomes of Synechocystis sp. PCC 6803 and Anabaena sp. PCC 7120 were analyzed for the respective predicted physical properties of each enzyme investigated here. Enzymatic activity was largely unaffected by nutritional mode, with the exception of glucokinase and PK whose activities were significantly elevated in heterotrophic cultures of Synechocystis sp. PCC 6803. PFK activity was insensitive to bacterial PFK-A (allosteric) effectors such as PEP, implying that Synechocystis PFK should be classified as a PFK-B (non-allosteric). Immunoblot and kinetic studies indicated that irrespective of nutritional mode, the Synechocystis PK corresponds to a PK-A (AMP activated) rather than PK-F (fructose-1,6-bisphosphate activated).     

Metabolic and molecular responses in Nile tilapia,Oreochromis niloticus during short and prolonged hypoxia

The strictly aquatic breathing Nile tilapia, Oreochromis niloticus is an extremely hypoxia-tolerant fish. To augment our understanding of the effects of hypoxia on anaerobic glycolysis in the Nile tilapia, we studied the effect of short-term for 1 day (trial 1) and long-term for 30 days (trial 2) hypoxia on a selected glycolytic enzymes activity and mRNA expression in liver and white muscle. The hypoxic oxygen concentrations used in the two trials were 2, 1, and 0.5 mg O₂ L^?1 for comparison with a control normoxic group 8 mg O₂ L^?1. The activity of phosphofructokinase (PFK), pyruvate kinase (PK), and lactate dehydrogenase (LDH) in liver and white muscle except liver LDH decreased in trial 1 and increased in trial 2. Assessments of mRNA levels in trial 1 revealed that PFK was downregulated and LDH was upregulated in liver and white muscle, while PK fluctuated between upregulation in liver and downregulation in white muscle. Meanwhile, PK and LDH were upregulated while PFK was similar to control values in both tissues in trial 2. Comet assay results demonstrated an increase in DNA damage that was directly proportional to increasing hypoxic concentrations. This damage was more pronounced in trial 1. This suggests that the Nile tilapia cope better with long-term hypoxic conditions, possibly as an adaptive response.     

Rat brain glycolysis regulation by estradiol-17 beta.

The effect of estradiol-17 beta on the activities of glycolytic enzymes from female rat brain was studied. The following enzymes were examined: hexokinase (HK, EC 2.7.1.1), phosphofructokinase (PFK, EC 2.7.1.11), aldolase (EC 4.1.2.13), glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.12), phosphoglycerate kinase (EC 2.7.2.3), phosphoglycerate mutase (EC 2.7.5.3), enolase (EC 4.2.1.11) and pyruvate kinase (PK, EC 2.7.1.40). The activities of HK (soluble and membrane-bound), PFK and PK were increased after 4 h of hormone treatment, while the others remained constant. The changes in activity were not seen in the presence of actinomycin D. The significant rise of the activities of the key glycolytic enzymes was also observed in the cell culture of mouse neuroblastoma C1300 treated with hormone. Only three of the studied isozymes, namely, HKII, B4 and K4 were found to be estradiol-sensitive for HK, PFK and PK, respectively. The results obtained suggest that rat brain glycolysis regulation by estradiol is carried out in neurons due to definite isozymes induction.     

Changes in activity of the regulatory glycolytic enzymes and of the pyruvate-dehydrogenase complex during the development of Xenopus laevis

In this study we investigated the variations of the maximal activities of the rate-controlling glycolytic enzymes (i.e., hexokinase, HK; phosphofructokinase, PFK; pyruvate kinase, PK) and of the pyruvate-dehydrogenase complex (PDHc) during the early embryogenesis of Xenopus laevis (from cleavage through hatching). All the enzymatic assays, using different coupled reactions, were performed spectrophotometrically on cytosolic and mitochondrial fractions. The maximal HK activity increases markedly from neurulation onwards, PFK activity presents a peak around gastrulation, PK activity remains relatively constant throughout the period studied and the highest PDHc activity is observed during cleavage. The specific activities display the same temporal pattern. Furthermore, in the sequence of reactions by which glucose is degraded to form acetyl-CoA, the maximal activities of PFK and PK are not limiting while those of HK and PDHc could be rate-limiting at relatively late developmental stages (hatching).