Alteration in liver pyruvate kinase protein and catalytic activity upon starvation and refeeding

Liver pyruvate kinase (L-type isozyme) was purified from the livers of rats fed a high carbohydrate, low protein diet for 4 days. The protein was homogeneous as judged by polyacrylamide-gel electrophoresis with and without added sodium dodecyl sulfate and as judged by high speed sedimentation and low speed equilibrium centrifugation. The specific activity of the purified protein was 190–220 international units (IU)/mg. A precipitating antiserum directed specifically against liver pyruvate kinase was obtained from rabbits and was used to determine the amount of liver pyruvate kinase protein present in the 80,000g supernatant fraction of rat liver homogenates in response to the dietary status of the animal. Rats maintained on a high carbohydrate, low protein diet for 4 days prior to sacrifice have at least 20 mg of precipitable liver pyruvate kinase protein per liver. Starvation of the animal results in a marked reduction in liver pyruvate kinase so that by 3 days of starvation less than 7 mg of liver pyruvate kinase protein per liver remains. Refeeding the animal a high carbohydrate, low protein diet results in a return of the liver pyruvate kinase protein to the prestarvation level of 20 mg per liver. The liver pyruvate kinase activity per liver varies in the same direction as does the liver pyruvate kinase protein but does not parallel the change in protein. Animals fed a high carbohydrate, low protein diet for 4 days have 60–70 IU/mg of liver pyruvate kinase protein whereas animals starved for periods exceeding 30 h have greater than 100 IU/mg of liver pyruvate kinase protein. Refeeding starved animals with a high carbohydrate, low protein diet initially causes a large increase in activity per milligram of liver pyruvate kinase protein followed by a return of this value to the prestarvation level. The observed rise in the ratio of activity per milligram of liver pyruvate kinase protein during starvation suggests a modification in the enzyme protein resulting either in an increase in the specific activity of the enzyme or in a decrease in the affinity of the enzyme for the antibody.     

The immunological properties of pyruvate kinase. II. The relationship of the human erythrocyte isozyme to the human liver isozymes.

We have examined the hypothesis that the human erythrocyte isozyme of pyruvate kinase (EC 2.7.1.40) is a hybrid of the two isozymes present in liver. Rabbit antiserum against purified human erythrocyte pyruvate kinase inactivates the erythrocyte isozyme and the major liver isozyme from human tissue but does not inactivate the minor liver isozyme. The electrophoretic mobilities of the erythrocyte and major liver isozymes are altered by anti-erythrocyte enzyme antibody while the mobility of the minor liver isozyme is unaffected. Gel diffusion analysis indicates cross-reactivity between the erythrocyte and major liver isozyme but no cross-reactivity with the minor liver isozyme. The hybrid hypothesis would predict cross-reactivity including changes in activity and mobility of all isozymes and we conclude, therefore that the hypothesis is incorrect.     

Long-term modulation of type L pyruvate kinase activity in young and mature rats

The regulation of type L pyruvate kinase concentrations in liver of young (35–45 days old) and adult (60–85 days old) rats starved and re-fed a 71% sucrose diet was investigated. Re-feeding is accompanied by an increase in the enzyme level in liver determined kinetically and immunologically. A constant ratio of kinetic activity to immunological activity was observed under all conditions examined, indicating that activity changes are the result of a regulation of synthesis or degradation and not an interconversion between kinetically active and inactive forms of the enzyme. Synthesis of pyruvate kinase was directly examined by using hepatocytes isolated from starved and re-fed rats. A stimulation of pyruvate kinase synthesis is observed on re-feeding. This increase in synthesis of pyruvate kinase is retained by the isolated hepatocyte for up to 7h in the absence of hormonal stimuli. Administration of glucagon (1μm) to the isolated hepatocytes had no influence on synthesis of pyruvate kinase and no evidence for a glucagon-directed degradation of the enzyme was found. Re-feeding the rat was followed by a transient increase in the synthesis of pyruvate kinase. The peak rate of synthesis was observed before a detectable increase in the enzyme concentration. After a rapid synthesis period, a new steady-state level of the enzyme was achieved and synthesis rates declined. The time course and magnitude for the response to the sucrose diet was dependent on the age of the rat. In young rats, an increase in pyruvate kinase synthesis is observed within 6h and peak synthesis occurs at 11h after re-feeding sucrose. The peak synthesis rate for pyruvate kinase for young rats represents approx. 1% of total protein synthesis. With adult rats, increased pyruvate kinase synthesis is not observed for 11h, with peak synthesis occurring at 24h after re-feeding. In the older rats, peak pyruvate kinase synthesis constitutes greater than 4% of total protein synthesis. Continued re-feeding of the adult rat beyond 24h is accompanied by a decline of pyruvate kinase synthesis to approx. 1.5% of total protein synthesis. The concentration of the enzyme, however, does not decline during this period, suggesting that control of pyruvate kinase degradation as well as synthesis occurs.     

Hepatic pyruvate kinase. Regulation by glucagon, cyclic adenosine 3'-5'-monophosphate, and insulin in the perfused rat liver.

A reversible interconversion of two kinetically distinct forms of hepatic pyruvate kinase regulated by glucagon and insulin is demonstrated in the perfused rat liver. The regulation does not involve the total enzyme content of the liver, but rather results in a modulation of the substrate dependence. The forms of pyruvate kinase in liver homogenates are distinguished by measurements of the ratio of the enzyme activity at a subsaturating concentration of P-enolpyruvate (1.3 mM) to the activity at a saturating concentration of this substrate (6.6 mM). A low ratio form of pyruvate kinase (ratio between 0.1 and 0.2) is obtained from livers perfused with 10(-7) M glucagon or 0.1 mM adenosine 3':5'-monophosphate (cyclic AMP). A high ratio form of the enzyme is obtained from livers perfused with no hormone (ratio = 0.35 to 0.45). The regulation of pyruvate kinase by glucagon and cyclic AMP occurs within 2 min following the hormone addition to the liver. Insulin (22 milliunits/ml) counteracts the inhibition of pyruvate kinase caused by 5 X 10(-11) M glucagon, but has only a slight influence on the enzyme properties in the absence of the hyperglycemic hormone. The low ratio form of pyruvate kinase obtained from livers perfused with glucagon or cyclic AMP is unstable in liver extracts and will revert to a high ratio form within 10 min at 37 degrees or within a few hours at 0 degrees. Pyruvate kinase is quantitatively precipitated from liver supernatants with 2.5 M ammonium sulfate. This precipitation stabilizes the enzyme and preserves the kinetically distinguishable forms. The kinetic properties of the two forms of rat hepatic pyruvate kinase are examined using ammonium sulfate precipitates from the perfused rat liver. At pH 7.5 the high ratio form of the enzyme has [S]0.5 = 1.6 +/- 0.2 mM P-enolpyruvate (n = 8). The low ratio form of enzyme from livers perfused with glucagon or cyclic AMP has [S]0.5 = 2.5 +/- 0.4 mM P-enolpyruvate (n = 8). The modification of pyruvate kinase induced by glucagon does not alter the dependence of the enzyme activity on ADP (Km is approximately 0.5 mM ADP for both forms of the enzyme). Both forms are allosterically modulated by fructose 1,6-bisphosphate, L-alanine, and ATP. The changes in the kinetic properties of hepatic pyruvate kinase which follow treating the perfused rat liver with glucagon or cyclic AMP are consistent with the changes observed in the enzyme properties upon phosphorylation in vitro by a clyclic AMP-stimulated protein kinase (Ljungstr?m, O., Hjelmquist, G. and Engstr?m, L. (1974) Biochim. Biophys. Acta 358, 289--298). However, other factors also influence the enzyme activity in a similar manner and it remains to be demonstrated that the regulation of hepatic pyruvate kinase by glucagon and cyclic AMP in vivo involes a phosphorylation.     

Age-related changes in activities of hepatic phosphofructokinase, pyruvate kinase and pyruvate dehydrogenase in liver and adipose tissue of the swiss albino mouse

The activities of phosphofructokinase, pyruvate kinase and pyruvate dehydrogenase were examined in liver as a function of age in Swiss albino mice. The hepatic activity of phosphofructokinase and total pyruvate dehydrogenase peaked in mice between 8 and 12 weeks of age and then decreased to a value that remained stable in mature animals older than 24 weeks of age. Yet, the activity of pyruvate kinase and pyruvate dehydrogenase in the active form in liver remained unchanged in mice up to 12 weeks of age. As mice matured, a progressive increase in the activity of both pyruvate kinase and the active form of pyruvate dehydrogenase in liver was observed while phosphofructokinase was unaltered. The pyruvate dehydrogenase complex, both total activity and the proportion of the enzyme in the active form, in the epididymal fat pad of the mouse showed no consistent age trend. The observed increase in the activity of both pyruvate kinase and the active form of pyruvate dehydrogenase should provide an augmented capacity for the generation of acetyl-CoA units for de novo fatty acid synthesis in livers of mature mice.     

Effects of hibernation on mitochondrial regulation and metabolic capacities in myocardium of painted turtle (Chrysemys picta)

Painted turtles hibernating during winter may endure long-term exposure to low temperature and anoxia. These two conditions may affect the aerobic capacity of a tissue and might be of particular importance to the cardiac muscle normally highly reliant on aerobic energy production. The present study addressed how hibernation affects respiratory characteristics of mitochondria in situ and the metabolic pattern of turtle myocardium. Painted turtles were acclimated to control (25 degrees C), cold (5 degrees C) normoxic and cold anoxic conditions. In saponin-skinned myocardial fibres, cold acclimation increased mitochondrial respiratory capacity and decreased apparent ADP-affinity. Concomitant anoxia did not affect this. Creatine increased the apparent ADP-affinity to similar values in the three acclimation groups, suggesting a functional coupling of creatine kinase to mitochondrial respiration. As to the metabolic pattern, cold acclimation decreased glycolytic capacity in terms of pyruvate kinase activity and increased lactate dehydrogenase (LHD) activity. Concomitant anoxia counteracted the cold-induced decrease in pyruvate kinase activity and increased creatine kinase activity. In conclusion, cold acclimation seems to increase aerobic and decrease anaerobic energy production capacity in painted turtle myocardium. Importantly, anoxia does not affect the mitochondrial functional integrity but seems to increase the capacity for anaerobic energy production and energy buffering.     

Effect of methylguanidine and guanidinosuccinic acid on pyruvate kinase activity in human red cells

An effect of methylguanidine and guanidinosuccinic acid on pyruvate kinase activity in human red cells was determined in vitro following a 3-hour incubation at 37 degrees C. The obtained results have shown that methylguanidine in the concentration of 1.8 x 10(-5) M/l inhibits pyruvate kinase activity by 20.8%. Pyruvate kinase activity was statistically significantly inhibited on addition of methylguanidine in the concentration of 5.4 x 10(-5) M/l whereas higher concentrations have no such an effect Guanidinesuccinic acid exerted similar but weaker effect on the activity of pyruvate kinase in human red cells. Mixture of methylguanidine (5.4 x 10(-5) m/l) and guanidinesuccinic acid (2.8 x 10(-5) M/l) does not affect pyruvate kinase activity in normal human red cells under identical experimental conditions.     

Phosphorylation of human erythrocyte pyruvate kinase by soluble cyclic-AMP-dependent protein kinases. Comparison with human liver L-type enzyme

Human red cell contain soluble adenosine-3',5'-phosphate-dependent protein kinases, which are able to phosphorylate the L' subunits of erythrocyte pyruvate kinase. Efficiency and maximum level of phosphorylation are very comparable in human liver and red cells. Phosphorylation of red cell pyruvate kinase results in the same kinetic modifications as for liver enzyme, namely a shift towards a 'T' allosteric state characterized by a decreased affinity for phosphoenolpyruvate and increased inhibition by the allosteric inhibitors ATP and alanine. In the course of red cell aging a small amount of partially proteolysed pyruvate kinase, devoid of the phosphorylatable site, appears; it resembles the subtilisin-treated L'4 enzyme and accounts for less than 20% of total pyruvate kinase subunits. Endogenous phosphorylation of pyruvate kinase from erythrocytes incubated in the presence of cyclic nucleotides produces the same kinetic modifications as phosphorylation in partially purified extract; this, however, does not change glucose consumption, lactate production and glycolytic intermediate concentrations of the incubated cells.     

Comparison of Isocitrate Dehydrogenase Activity, Pyruvate Kinase Activity, and Polyphenol Content in Physiologically Different Pine Callus Tissue

During the growth of callus tissue of slash pine (Pinus elliottil Engelm.) several physiologically different types of tissue can be observed, often within the same culture. Different tissues were selected, based on color appearance, and used to determine isocitrate dehydrogenase and pyruvate kinase activity, and total polyphenol content. Isocitrate dehydrogenase and pyruvate kinase activity in yellow tissue was 3- to 5-fold greater than in brown tissue, whereas the polyphenol content in yellow tissue was approximately 5-fold less than in brown tissue. Dark brown callus, which also contained large amounts of polyphenols, did not have detectable enzyme activity. The differences in optimal concentrations of substrate and cofactors for the isocitrate dehydrogenase and pyruvate kinase reactions in yellow and brown tissues were very minor and therefore cannot account for the 3- to 5-fold difference in enzyme activity between these tissues. Also, the addition of brown or dark-brown tissue extract to the yellow tissue extract did not inhibit isocitrate dehydrogenase or pyruvate kinase activity in the yellow tissue extract.     

Biosynthesis of rat liver pyruvate kinase. Measurement of enzyme lifetime and the rate of synthesis at weaning.

Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis of immunoprecipitates of liver cytosol with anti-(L-type pyruvate kinase) serum revealed proteins of mol.wt. 56 000 and 42 000 in addition to the heavy and light chains. The ratio of the 56 000 mol.wt. to the 42 000 mol.wt. protein increased under dietary conditions that resulted in an increase in the apparent specific activity of hepatic pyruvate kinase. The 42 000 mol.wt. protein was removed from immunoprecipitates if the liver cytosol was partially purified by pH precipitation and (NH4)2SO4 fractionation before addition of the antiserum. This technique may be used to analyse the formation of pure L-type pyruvate kinase in liver. By using H14CO3-labelling, the t1/2 of L-type pyruvate kinase was estimated as 75 +/- 1.7 h in post-weaned high-carbohydrate-diet-fed rats. Before weaning there was little immunoreactive pyruvate kinase in rat liver cytosol. Induction began between 6 and 24 h after weaning and reached a maximum value 120 h after weaning. When clearly enhanced total pyruvate kinase activity was first observed at 24 h post-weaning, the apparent specific activity of hepatic pyruvate kinase was considerably lower than the specific activity of the pure isolated enzyme. When the induction of L-type pyruvate kinase was monitored by the incorporation of L-[4,5-3H]leucine, the maximum rate of synthesis occurred 24--48 h after weaning. After this period synthesis declined, indicating a relatively slow turnover of the enzyme once the enzyme concentration was established in the liver.     

An allele (Pk-1 b ) from wild-caught mice that affects the activity and kinetics of erythrocyte and liver pyruvate kinase

A true breeding strain was made from a wild-caught mouse with low erythrocyte pyruvate kinase (E.C. 2.7.1.40) activity. This variation showed additive inheritance and segregated as an allele at a single locus (Pk-1 ^b). Mice homozygous for the reduced blood pyruvate kinase activity cosegregated for reduced liver activity. In both these tissues the variant enzyme had a lowered heat stability and reduced K _m values for ADP. An increased stimulation by FDP was also detected in the liver pyruvate kinase. No difference in the isoelectric point of the variant enzyme in either erythrocyte or liver was observed when compared with the enzyme from C57BL mice (Pk-1 ^a/Pk-1 ^a). It is concluded that Pk-1 is the structural gene for the erythrocyte and the major liver pyruvate kinase. No other tissue pyruvate kinase showed altered characteristics.This work was supported by a Medical Research Council grant.     

Evidence for calcium enhanced phosphorylation of pyruvate kinase by pancreatic islets

Summary Pancreatic islet cytosol contains a calcium-calmodulin dependent protein kinase that can mediate the phosphorylation of an endogenous protein that has an Mr of 57 000, as well as exogenous muscle pyruvate kinase (subunit Mr, 57000). EGTA and trifluoperazine decreased the phosphorylation. Alkaline inactivation of pyruvate kinase made it a better substrate for the kinase. As in rat islet cytosol, rabbit islet cytosol catalyzed the phosphorylation of a 57 000 Mr protein in the presence of calcium and calmodulin. This phosphoprotein was immunoprecipitated with anti-pyruvate kinase antibody. This is consistent with the idea that the 57 000 Mr phosphoprotein in islet cytosol is the subunit of pyruvate kinase. The paper following this paper shows that the kinetic and immunologic properties of the islet pyruvae kinase indicate it is the M₂ isoenzyme and that its phosphorylation does not affect its catalytic activity.     

Pancreatic islets contain the M2 isoenzyme of pyruvate kinase its phosphorylation has no effect on enzyme activity

To determine which of the major isoenzymes of pyruvate kinase pancreatic islet pyruvate kinase most resembled, it was compared to pyruvate kinase from other tissues in kinetic and immunologic studies. The pattern of activation by fructose bisphosphate and the patterns of inhibition by alanine and phenylalanine were most similar to those of the M₂ isoenzyme from kidney and were dissimilar to those of the isoenzymes from skeletal muscle (type M₁) and liver (type L). The islet pyruvate kinase was inhibited by anti-M₁ pyruvate kinase serum (which crossreacts with the M₂ isoenzyme), but not by anti-L pyruvate kinase. These results are most consistent with islets possessing predominantly, if not exclusively, the M₂ isoenzyme of pyruvate kinase. We previously showed that rat pancreatic islet cytosol contains protein kinases that can catalyze a calcium-activated phosphorylation of an endogenous peptide that has properties, such as subunit molecular weight and isoelectric pH, that are identical to those of the M₂ and M, isoenzymes of pyruvate kinase, and that islet cytosol can catalyze phosphorylation of muscle pyruvate kinase. In the present study it was shown that incubating islet cytosol with ATP under conditions known to permit phosphorylation and inhibition of liver pyruvate kinase did not affect the islet pyruvate kinase activity. It is concluded that phosphorylation of the islet pyruvate kinase has no immediate effect on enzyme activity.Abbreviations EGTA ethylene glycos his (-aminoethyl ether)-N,N,NN-tetraacetic acid - Hepes 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid     

A comparison of the regulation of pyruvate dehydrogenase in mitochondria from rat brain and liver.

The total activity of pyruvate dehydrogenase in mitochondria isolated from rat brain and liver was 53.5 and 14.2nmol/min per mg of protein respectively. Pyruvate dehydrogenase in liver mitochondria incubated for 4 min at 37 degrees C with no additions was 30% in the active form and this activity increased with longer incubations until it was completely in the active form after 20 min. Brain mitochondrial pyruvate dehydrogenase activity was initially high and did not increase with addition of Mg2+ plus Ca2+ or partially purified pyruvate dehydrogenase phosphatase or with longer incubations. The proportion of pyruvate dehydrogenase in the active form in both brain and liver mitochondria changed inversely with changes in mitochondrial energy charge, whereas total pyruvate dehydrogenase did not change. The chelators citrate, isocitrate, EDTA, ethanedioxybis(ethylamine)tetra-acetic acid and Ruthenium Red each lowered pyruvate dehydrogenase activity in brain mitochondria, but only citrate and isocitrate did so in liver mitochondria. These chelators did not affect the energy charge of the mitochondria. Mg2+ plus Ca2+ reversed the pyruvate dehydrogenase inactivation in liver, but not brain, mitochondria. The regulation of the activation-inactivation of pyruvate dehydrogenase in mitochondria from rat brain and liver with respect to energy charge is similar and may be at least partially regulated by this parameter, and the effects of chelators differ in the two types of mitochondria.     

Dexamethasone effects on liver pyruvate kinase

Dexamethasone in the medium perfusin isolated rabbit livers caused a fast-acting and reversible effect on liver pyruvate kinase. The effect was to lower th assayable V activity (units/g tissue) without changing the concentration (nmol/g enzyme protein). In effect, glucocorticoid lowered the specific activity (units/nmol of enzyme) by direct action on liver. The effect on liver pyruvate kinase is mediated by a relatively stable alteration; 30 min after perfusate (with steroid) was replaced by perfusate (without steroid), the effect remained strongly evident.     

Characterization of an Escherichia coli K12 mutant that is sensitive to chlorate when grown aerobically.

1. The ;initial activity' of the pyruvate dehydrogenase enzyme complex in whole tissue or mitochondrial extracts of lactating rat mammary glands was greatly decreased by 24 or 48h starvation of the rats. Injection of insulin and glucose into starved rats 60min before removal of the glands abolished this difference in ;initial activities'. 2. The ;total activity' of the enzyme complex in such extracts was revealed by incubation in the presence of free Mg(2+) and Ca(2+) ions (more than 10 and 0.1mm respectively) and a crude preparation of pig heart pyruvate dehydrogenase phosphatase. Starvation did not alter this ;total activity'. It is assumed that the decline in ;initial activity' of the enzyme complex derived from the glands of starved animals was due to increased phosphorylation of its alpha-subunit by intrinsic pyruvate dehydrogenase kinase. 3. Starvation led to an increase in intrinsic pyruvate dehydrogenase kinase activity in both whole tissue and mitochondrial extracts. Injection of insulin into starved animals 30min before removal of the lactating mammary glands abolished the increase in pyruvate dehydrogenase kinase activity in whole-tissue extracts. 4. Pyruvate (1mm) prevented ATP-induced inactivation of the enzyme complex in mitochondrial extracts from glands of fed animals. In similar extracts from starved animals pyruvate was ineffective. 5. Starvation led to a decline in activity of pyruvate dehydrogenase phosphatase in mitochondrial extracts, but not in whole-tissue extracts. 6. These changes in activity of the intrinsic kinase and phosphatase of the pyruvate dehydrogenase complex of lactating rat mammary gland are not explicable by current theories of regulation of the complex.     

A requirement for DNA synthesis in foetal hepatocyte differentiation

In hepatocyte cultures derived from 15-day-old foetal rats, the appearance of the liver (L) form of pyruvate kinase is blocked when cytosine arabinoside is added on the 2nd day of culture. When added on the 3rd day of culture, the inhibitor of DNA synthesis does not prevent the appearance of the enzyme. If cytosine arabinoside is added on the 2nd day of culture and removed on the 4th day, the enzyme is detected by the 6th day of culture. The specificity of the action of cytosine arabinoside for the L form of pyruvate kinase is in contrast with the lack of effect observed on total protein synthesis and the activity of the embryonic (M2) form of the enzyme.     

Inactivation of pyruvate kinase type M2 from chicken liver by phosphorylation, catalyzed by a cAMP-independent protein kinase.

A cAMP-independent protein kinase from chicken liver phosphorylated and inactivated pyruvate kinase type M2 from the same tissue. Complete inactivation was reached when 4 mol of phosphate were incorporated/mol of tetrameric pyruvate kinase. The protein kinase bound with high affinity to pyruvate kinase type M2 (Km value for pyruvate kinase = 6 X 10(-10)M; it phosphorylated phosvitin and casein but not histones, ATP and GTP were substrates. The differences between the properties of this protein kinase in the interconversion of pyruvate kinase and that described previously are discussed.     

Control of gluconeogenesis in rat liver cells. I. Kinetics of the individual enzymes and the effect of glucagon

Control of gluconeogenesis from lactate was studied by titrating rat liver cells with lactate and pyruvate in a ratio of 10:1 in a perifusion system. At different steady states of glucose formation, the concentration of key gluconeogenic intermediates was measured and plotted against gluconeogenic flux (J glucose). Complete saturation was observed only in the plot relating J glucose to the extracellular pyruvate concentration. Measurement of pyruvate distribution in the cell showed that the mitochondrial pyruvate translocator operates close to equilibrium at high lactate and pyruvate concentrations. It can therefore be concluded that pyruvate carboxylase limits maximal gluconeogenic flux. Addition of glucagon did not cause a shift in the plots relating J glucose to glucose 6-phosphate, dihydroxyacetone phosphate, 3-phosphoglycerate, and phosphoenolpyruvate. It can thus be concluded that glucagon does not affect the kinetic parameters of the enzymes involved in the conversion of phosphoenolpyruvate to glucose. Addition of glucagon led to a shift in the curves relating J glucose to the concentration of cytosolic oxalacetate and extracellular pyruvate. The shift in the curve relating J glucose to oxalacetate is due to glucagon-induced inhibition of pyruvate kinase. The stimulation of gluconeogenesis by glucagon can be accounted for almost completely by inhibition of pyruvate kinase. There was almost no stimulation by glucagon of pyruvate carboxylation. In the absence of glucagon, control on gluconeogenesis from lactate is distributed among different steps including pyruvate carboxylase and pyruvate kinase. Assuming that in the presence of glucagon all pyruvate kinase flux is inhibited, the control of gluconeogenesis in the presence of the hormone is confined exclusively to pyruvate carboxylase.     

The dual activity of pyruvate kinase type M2 from chromatin extracts of neoplastic cells

Pyruvate kinase type M(2) from Morris hepatoma 7777 tumour cell nuclei and cytosol, in contrast to types L and M(2) from nuclei and cytosol of normal rat liver, shows the histone H(1) kinase activity. Moreover, in the presence of L-cysteine and without ADP it converts 2-phosphoenolpyruvate (PEP) to pyruvate while in the presence of L-arginine or L-histidine does not. L-Cysteine markedly stimulates the activity of histone H(1) kinase transferring a phosphate group from PEP to, as results suggested, the epsilon -amino group of L-lysine of histone H(1). This, L-cysteine which is known to inhibit the activity of pyruvate kinase type M(2) from neoplastic cells transfering a phosphate from PEP to ADP, can act as a control factor champing the direction of enzymatic reaction in cancer cells.