Muscle-Type 6-Phosphofructo-1-kinase and Aldolase Associate Conferring Catalytic Advantages for Both Enzymes

Laboratório de Oncobiologia Molecular (LabOMol), Departamento de Fármacos, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ, Brazil.
International Union of Biochemistry and Molecular Biology Life (Impact Factor: 3.14). 06/2011; 63(6):435-45. DOI: 10.1002/iub.464
Source: PubMed


6-Phosphofructo-1-kinase (PFK) and aldolase are two sequential glycolytic enzymes that associate forming heterotetramers containing a dimer of each enzyme. Although free PFK dimers present a negligible activity, once associated to aldolase these dimers are as active as the fully active tetrameric conformation of the enzyme. Here we show that aldolase-associated PFK dimers are not inhibited by clotrimazole, an antifungal azole derivative proposed as an antineoplastic drug due to its inhibitory effects on PFK. In the presence of aldolase, PFK is not modulated by its allosteric activators, ADP and fructose-2,6-bisphosphate, but is still inhibited by citrate and lactate. The association between the two enzymes also results on the twofold stimulation of aldolase maximal velocity and affinity for its substrate. These results suggest that the association between PFK and aldolase confers catalytic advantage for both enzymes and may contribute to the channeling of the glycolytic metabolism.

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Available from: Mauro Sola-Penna, May 07, 2014
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    • "Skeletal muscle aldolase has also been shown to interact and decrease the inactivation of the enzyme phosphofructokinase [15,16]. This association not only alters the allosteric regulation of phosphofructokinase but also increases the activity of aldolase by approximately 2-fold [17], which may provide an advantage for channelling substrates through the glycolytic pathway. However, skeletal muscle and liver express different aldolase isoforms, aldolase B being preferentially expressed in liver and isoform A in the skeletal muscle [18]. "
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    ABSTRACT: Previous studies have demonstrated that glucose disposal is increased in the Fyn knockout (FynKO) mice due to increased insulin sensitivity. FynKO mice also display fasting hypoglycaemia despite decreased insulin levels, which suggested that hepatic glucose production was unable to compensate for the increased basal glucose utilization. The present study investigates the basis for the reduction in plasma glucose levels and the reduced ability for the liver to produce glucose in response to gluconeogenic substrates. FynKO mice had a 5-fold reduction in phosphoenolpyruvate carboxykinase (PEPCK) gene and protein expression and a marked reduction in pyruvate, pyruvate/lactate-stimulated glucose output. Remarkably, de novo glucose production was also blunted using gluconeogenic substrates that bypass the PEPCK step. Impaired conversion of glycerol to glucose was observed in both glycerol tolerance test and determination of the conversion of (13)C-glycerol to glucose in the fasted state. α-glycerol phosphate levels were reduced but glycerol kinase protein expression levels were not changed. Fructose-driven glucose production was also diminished without alteration of fructokinase expression levels. The normal levels of dihydroxyacetone phosphate and glyceraldehyde-3-phosphate observed in the FynKO liver extracts suggested normal triose kinase function. Fructose-bisphosphate aldolase (aldolase) mRNA or protein levels were normal in the Fyn-deficient livers, however, there was a large reduction in liver fructose-6-phosphate (30-fold) and fructose-1,6-bisphosphate (7-fold) levels as well as a reduction in glucose-6-phosphate (2-fold) levels. These data suggest a mechanistic defect in the allosteric regulation of aldolase activity.
    PLoS ONE 11/2013; 8(11):e81866. DOI:10.1371/journal.pone.0081866 · 3.23 Impact Factor
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    ABSTRACT: The present work describes the effects of metformin on hexokinase (HK) and phosphofructokinase (PFK) activities and localization in different tissues from streptozotocin-induced diabetic mice. Diabetic mice present lower HK and PFK activities (50%) in skeletal muscle, liver and adipose tissue, as compared with control (P<0.05). Treatment with 250 mg/kg metformin reverses this pattern of enzyme inhibition with concomitant reversal of hyperglycemia and hypolactacidemia. Furthermore, the treatment increases the cytoskeleton-associated PFK activity in skeletal muscle; this activity has been described as an important mechanism for the enzyme activation. This effect might be due to the increased phosphorylation of serine residues in the enzyme, a modification which has been described to increase the interaction of PFK with f-actin. The current work supports the hypothesis that metformin hypoglycemic effects involve the activation of glycolysis through its regulatory enzymes, which may be potential targets for the development of new hypoglycemic drugs.
    Archives of Biochemistry and Biophysics 04/2010; 496(1):53-60. DOI:10.1016/ · 3.02 Impact Factor
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    ABSTRACT: Diabetes mellitus is characterized by hyperglycemia and its associated complications, including cardiomyopathy. Metformin, in addition to lowering blood glucose levels, provides cardioprotection for diabetic subjects. Glycolysis is essential to cardiac metabolism and its reduction may contribute to diabetic cardiomyopathy. Hexokinase (HK) and phosphofructokinase (PFK), rate-limiting enzymes of glycolysis, are downregulated in cardiac muscle from diabetic subjects, playing a central role on the decreased glucose utilization in the heart of diabetic subjects. Thus, the aim of this study was to determine whether metformin modulates heart HK and PFK from diabetic mice. Diabetes was induced by streptozotocin injection on male Swiss mice, which were treated for three consecutive days with 250 mg/kg metformin before evaluating HK and PFK activity, expression, and intracellular distribution on the heart of these subjects. We show that metformin abrogates the downregulation of HK and PFK in the heart of streptozotocin-induced diabetic mice. This effect is not correlated to alteration on the enzymes' transcription and expression. However, the intracellular distribution of both enzymes is altered in diabetic hearts that show increased activity of the soluble fraction when compared to the particulate fraction. Moreover, this pattern is reversed upon the treatment with metformin, which is correlated with the effects of the drug on the enzymes activity. Altogether, our results support evidences that metformin alter the intracellular localization of HK and PFK augmenting glucose utilization by diabetic hearts and, thus, conferring cardiac protection to diabetic subjects.
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