Isoenzymes of phosphofructokinase in the rat. Demonstration of the three non-identical subunits by biochemical, immunochemical and kinetic studies

Vora, Shobhana; Oskam, R.; Staal, Gerard E.J.

doi:10.1042/bj2290333

Cited by 41 publications

(32 citation statements)

References 27 publications

(55 reference statements)

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“…Taking the mitochondrial to cytoplasmic citrate ratio into account, estimated Cu− cerebellar cytoplasmic citrate would be approximately 15 μM (48). This citrate concentration, while inhibiting rat brain PFK2 activity by more than 80%, would have little impact on rat brain PFK1 activity (40, 49). Higher G6P levels in Cu− tissue may also be due to a potential rise in NADPH/NADP + , which has not been evaluated in Cu− cerebellum.…”

Section: Discussionmentioning

confidence: 99%

Fructose-2,6-Bisphosphate Is Lower in Copper Deficient Rat Cerebellum Despite Higher Content of Phosphorylated AMP-Activated Protein Kinase

Gybina

Prohaska

2008

Exp Biol Med (Maywood)

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Limitation in copper (Cu) leads to pathophysiology in developing brain. Cu deficiency impairs brain mitochondria and results in high brain lactate suggesting augmented anaerobic glycolysis. AMP activated protein kinase (AMPK) is a cellular energy "master-switch" that is thought to augment glycolysis through phosphorylation and activation phosphofructokinase 2 (PFK2) resulting in increases of the glycolytic stimulator fructose-2,6-bisphosphate (F2,6BP). Previously, Cu deficiency has been shown to augment cerebellar AMPK activation. Cerebella of Cu-adequate (Cu+) and Cudeficient (Cu−) rat pups were assessed to evaluate if AMPK activation in Cu− cerebella functioned to enhance PFK2 activation and increase F2,BP concentration. Higher levels of pAMPK were detected in Cu− cerebella. However, PFK2 activity, mRNA, and protein abundance were not affected by Cu deficiency. Surprisingly, F2,6BP levels were markedly lower in Cu− cerebella. Lower F2,6BP may be due to inhibition of PFK2 by citrate, as citrate concentration was significantly higher in Cu − cerebella. Data suggest AMPK activation in Cu− cerebellum does not augment glycolysis through a PFK2 mechanism. Furthermore, other metabolite data suggest that glycolysis may actually be blunted, since levels of glucose and glucose-6-phosphate were higher in Cu− cerebella than controls.

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Section: Discussionmentioning

confidence: 99%

Fructose-2,6-Bisphosphate Is Lower in Copper Deficient Rat Cerebellum Despite Higher Content of Phosphorylated AMP-Activated Protein Kinase

Gybina

Prohaska

2008

Exp Biol Med (Maywood)

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“…All three isoforms of PFK are activated by F6P and F26BP [12], but only PFKM and PFKL are activated by F16BP [13]–[15]. PFKFB is a bifunctional enzyme whose kinase and bisphosphatase domains catalyze the formation and hydrolysis reaction of F26BP, respectively [9], [16].…”

Section: Introductionmentioning

confidence: 99%

Bistability in Glycolysis Pathway as a Physiological Switch in Energy Metabolism

et al. 2014

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The flux of glycolysis is tightly controlled by feed-back and feed-forward allosteric regulations to maintain the body's glucose homeostasis and to respond to cell's growth and energetic needs. Using a mathematical model based on reported mechanisms for the allosteric regulations of the enzymes, we demonstrate that glycolysis exhibits multiple steady state behavior segregating glucose metabolism into high flux and low flux states. Two regulatory loops centering on phosphofructokinase and on pyruvate kinase each gives rise to the bistable behavior, and together impose more complex flux control. Steady state multiplicity endows glycolysis with a robust switch to transit between the two flux states. Under physiological glucose concentrations the glycolysis flux does not move between the states easily without an external stimulus such as hormonal, signaling or oncogenic cues. Distinct combination of isozymes in glycolysis gives different cell types the versatility in their response to different biosynthetic and energetic needs. Insights from the switch behavior of glycolysis may reveal new means of metabolic intervention in the treatment of cancer and other metabolic disorders through suppression of glycolysis.

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“…All three isoenzymes are strongly inhibited by citrate, with IC 50 values of 0.08, 0.13 and 0.18 mM for brain (platelet), muscle, and liver PFK1, respectively [17]. All human PFK1 isoforms are also reported to be intensely inhibited by ATP concentrations higher than 0.05 mM, yet (F-2,6-BP) can antagonise the negative effects of ATP to some extent [18].…”

Section: Introductionmentioning

confidence: 99%

Posttranslational Modification of 6-phosphofructo-1-kinase as an Important Feature of Cancer Metabolism

2011

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BackgroundHuman cancers consume larger amounts of glucose compared to normal tissues with most being converted and excreted as lactate despite abundant oxygen availability (Warburg effect). The underlying higher rate of glycolysis is therefore at the root of tumor formation and growth. Normal control of glycolytic allosteric enzymes appears impaired in tumors; however, the phenomenon has not been fully resolved.Methodology/Principal FindingsIn the present paper, we show evidence that the native 85-kDa 6-phosphofructo-1-kinase (PFK1), a key regulatory enzyme of glycolysis that is normally under the control of feedback inhibition, undergoes posttranslational modification. After proteolytic cleavage of the C-terminal portion of the enzyme, an active, shorter 47-kDa fragment was formed that was insensitive to citrate and ATP inhibition. In tumorigenic cell lines, only the short fragments but not the native 85-kDa PFK1 were detected by immunoblotting. Similar fragments were detected also in a tumor tissue that developed in mice after the subcutaneous infection with tumorigenic B16-F10 cells. Based on limited proteolytic digestion of the rabbit muscle PFK-M, an active citrate inhibition-resistant shorter form was obtained, indicating that a single posttranslational modification step was possible. The exact molecular masses of the active shorter PFK1 fragments were determined by inserting the truncated genes constructed from human muscle PFK1 cDNA into a pfk null E. coli strain. Two E. coli transformants encoding for the modified PFK1s of 45,551 Da and 47,835 Da grew in glucose medium. The insertion of modified truncated human pfkM genes also stimulated glucose consumption and lactate excretion in stable transfectants of non-tumorigenic human HEK cell, suggesting the important role of shorter PFK1 fragments in enhancing glycolytic flux.Conclusions/SignificancePosttranslational modification of PFK1 enzyme might be the pivotal factor of deregulated glycolytic flux in tumors that in combination with altered signaling mechanisms essentially supports fast proliferation of cancer cells.

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Isoenzymes of phosphofructokinase in the rat. Demonstration of the three non-identical subunits by biochemical, immunochemical and kinetic studies

Cited by 41 publications

References 27 publications

Fructose-2,6-Bisphosphate Is Lower in Copper Deficient Rat Cerebellum Despite Higher Content of Phosphorylated AMP-Activated Protein Kinase

Fructose-2,6-Bisphosphate Is Lower in Copper Deficient Rat Cerebellum Despite Higher Content of Phosphorylated AMP-Activated Protein Kinase

Bistability in Glycolysis Pathway as a Physiological Switch in Energy Metabolism

Posttranslational Modification of 6-phosphofructo-1-kinase as an Important Feature of Cancer Metabolism

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