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. 1992 Aug 1;285(Pt 3):767–771. doi: 10.1042/bj2850767

Physiological concentrations of 2-oxoglutarate regulate the activity of phosphoenolpyruvate carboxykinase in liver.

M A Titheradge 1, R A Picking 1, R C Haynes Jr 1
PMCID: PMC1132861  PMID: 1497614

Abstract

2-Oxoglutarate was found to inhibit purified rat liver phosphoenolpyruvate carboxykinase when the assay was performed in the direction of either phosphoenolpyruvate or oxaloacetate synthesis. The inhibition was competitive with respect to oxaloacetate or phosphoenolpyruvate, the Ki values being 0.32 +/- 0.04 mM 0.63 +/- 0.19 mM respectively. 2-Oxoglutarate inhibited non-competitively when tested against GTP or Mn2+. The reported cytosolic concentrations of 2-oxoglutarate in rat hepatocytes are such that the enzyme is likely to be significantly inhibited under basal conditions. The cytosolic concentration of 2-oxoglutarate is known to fall precipitously under the influence of glucagon and other hormones that stimulate gluconeogenesis, and it is suggested that the hormone-induced decrease in 2-oxoglutarate content would alleviate the inhibition of phosphoenolpyruvate carboxykinase and stimulate flux from oxaloacetate to phosphoenolpyruvate. The implications of this finding to the rationalization of the role of pyruvate kinase in the stimulation of gluconeogenesis in the fasted state are discussed.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Ballard F. J., Hanson R. W. Purification of phosphoenolpyruvate carboxykinase from the cytosol fraction of rat liver and the immunochemical demonstration of differences between this enzyme and the mitochondrial phosphoenolpyruvate carboxykinase. J Biol Chem. 1969 Oct 25;244(20):5625–5630. [PubMed] [Google Scholar]
  2. Bentle L. A., Lardy H. A. Interaction of anions and divalent metal ions with phosphoenolpyruvate carboxykinase. J Biol Chem. 1976 May 25;251(10):2916–2921. [PubMed] [Google Scholar]
  3. Blair J. B., James M. E., Foster J. L. Adrenergic control of glycolysis and pyruvate kinase activity in hepatocytes from young and old rats. J Biol Chem. 1979 Aug 25;254(16):7585–7590. [PubMed] [Google Scholar]
  4. Brinkworth R. I., Hanson R. W., Fullin F. A., Schramm V. L. Mn2+-sensitive and -insensitive forms of phosphoenolpyruvate carboxykinase (GTP). J Biol Chem. 1981 Nov 10;256(21):10795–10802. [PubMed] [Google Scholar]
  5. Chan T. M., Exton J. H. Studies on alpha-adrenergic activation of hepatic glucose output. J Biol Chem. 1978 Sep 25;253(18):6393–6400. [PubMed] [Google Scholar]
  6. DIXON M. The determination of enzyme inhibitor constants. Biochem J. 1953 Aug;55(1):170–171. doi: 10.1042/bj0550170. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Eisenstein A. B., Strack I. Effects of glucagon on carbohydrate synthesis and enzyme activity in rat liver. Endocrinology. 1968 Dec;83(6):1337–1348. doi: 10.1210/endo-83-6-1337. [DOI] [PubMed] [Google Scholar]
  8. Exton J. H., Park C. R. Control of gluconeogenesis in liver. 3. Effects of L-lactate, pyruvate, fructose, glucagon, epinephrine, and adenosine 3',5'-monophosphate on gluconeogenic intermediates in the perfused rat liver. J Biol Chem. 1969 Mar 25;244(6):1424–1433. [PubMed] [Google Scholar]
  9. Feliú J. E., Hue L., Hers H. G. Hormonal control of pyruvate kinase activity and of gluconeogenesis in isolated hepatocytes. Proc Natl Acad Sci U S A. 1976 Aug;73(8):2762–2766. doi: 10.1073/pnas.73.8.2762. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Foster D. O., Lardy H. A., Ray P. D., Johnston J. B. Alteration of rat liver phosphoenolpyruvate carboxykinase activity by L-tryptophan in vivo and metals in vitro. Biochemistry. 1967 Jul;6(7):2120–2128. doi: 10.1021/bi00859a033. [DOI] [PubMed] [Google Scholar]
  11. Garrison J. C., Borland M. K., Florio V. A., Twible D. A. The role of calcium ion as a mediator of the effects of angiotensin II, catecholamines, and vasopressin on the phosphorylation and activity of enzymes in isolated hepatocytes. J Biol Chem. 1979 Aug 10;254(15):7147–7156. [PubMed] [Google Scholar]
  12. Garrison J. C. The effects of glucagon, catecholamines, and the calcium ionophore A23187 on the phosphorylation of rat hepatocyte cytosolic proteins. J Biol Chem. 1978 Oct 10;253(19):7091–7100. [PubMed] [Google Scholar]
  13. Garrison J. C., Wagner J. D. Glucagon and the Ca2+-linked hormones angiotensin II, norepinephrine, and vasopressin stimulate the phosphorylation of distinct substrates in intact hepatocytes. J Biol Chem. 1982 Nov 10;257(21):13135–13143. [PubMed] [Google Scholar]
  14. Groen A. K., Vervoorn R. C., Van der Meer R., Tager J. M. Control of gluconeogenesis in rat liver cells. I. Kinetics of the individual enzymes and the effect of glucagon. J Biol Chem. 1983 Dec 10;258(23):14346–14353. [PubMed] [Google Scholar]
  15. HOLTEN D. D., NORDLIE R. C. COMPARATIVE STUDIES OF CATALYTIC PROPERTIES OF GUINEA PIG LIVER INTRA- AND EXTRAMITOCHONDRIAL PHOSPHOENOLPYRUVATE CARBOXYKINASES. Biochemistry. 1965 Apr;4:723–731. doi: 10.1021/bi00880a018. [DOI] [PubMed] [Google Scholar]
  16. Haynes R. C., Jr, Picking R. A. The role of inhibition of pyruvate kinase in the stimulation of gluconeogenesis by glucagon: a reevaluation. Arch Biochem Biophys. 1990 Nov 15;283(1):51–59. doi: 10.1016/0003-9861(90)90611-2. [DOI] [PubMed] [Google Scholar]
  17. Hue L., Felíu J. E., Hers H. G. Control of gluconeogenesis and of enzymes of glycogen metabolism in isolated rat hepatocytes. A parallel study of the effect of phenylephrine and of glucagon. Biochem J. 1978 Dec 15;176(3):791–797. doi: 10.1042/bj1760791. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Häussinger D., Sies H. Effect of phenylephrine on glutamate and glutamine metabolism in isolated perfused rat liver. Biochem J. 1984 Aug 1;221(3):651–658. doi: 10.1042/bj2210651. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Jomain-Baum M., Schramm V. L., Hanson R. W. Mechanism of 3-mercaptopicolinic acid inhibition of hepatic phosphoenolpyruvate carboxykinase (GTP). J Biol Chem. 1976 Jan 10;251(1):37–44. [PubMed] [Google Scholar]
  20. Jomain-Baum M., Schramm V. L. Kinetic mechanism of phosphoenolpyruvate carboxykinase (GTP) from rat liver cytosol. Product inhibition, isotope exchange at equilibrium, and partial reactions. J Biol Chem. 1978 May 25;253(10):3648–3659. [PubMed] [Google Scholar]
  21. Karczmarewicz E., Matyaszczyk M., Vorbrodt Z., Lorenc R. Activation of liver cytosol phosphoenolpyruvate carboxykinase by Ca2+ through intracellular redistribution of Mn2+. Eur J Biochem. 1985 Sep 16;151(3):561–565. doi: 10.1111/j.1432-1033.1985.tb09140.x. [DOI] [PubMed] [Google Scholar]
  22. McCormack J. G. Studies on the activation of rat liver pyruvate dehydrogenase and 2-oxoglutarate dehydrogenase by adrenaline and glucagon. Role of increases in intramitochondrial Ca2+ concentration. Biochem J. 1985 Nov 1;231(3):597–608. doi: 10.1042/bj2310597. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Merryfield M. L., Lardy H. A. Ca2+-mediated activation of phosphoenolpyruvate carboxykinase occurs via release of Fe2+ from rat liver mitochondria. J Biol Chem. 1982 Apr 10;257(7):3628–3635. [PubMed] [Google Scholar]
  24. Meyuhas O., Boshwitz C., Reshef L. Phosphoenolpyruvate carboxylase decarboxylation catalyzed reaction in cytosol of rat adipose tissue. Biochim Biophys Acta. 1971 Oct;250(1):224–237. doi: 10.1016/0005-2744(71)90138-0. [DOI] [PubMed] [Google Scholar]
  25. Ochs R. S. Glutamine metabolism of isolated rat hepatocytes. Evidence for catecholamine activation of alpha-ketoglutarate dehydrogenase. J Biol Chem. 1984 Nov 10;259(21):13004–13010. [PubMed] [Google Scholar]
  26. Ochs R. S., Lardy H. A. Catecholamine stimulation of hepatic gluconeogenesis at the site between pyruvate and phosphoenolpyruvate. J Biol Chem. 1983 Aug 25;258(16):9956–9962. [PubMed] [Google Scholar]
  27. Philippidis H., Hanson R. W., Reshef L., Hopgood M. F., Ballard F. J. The initial synthesis of proteins during development. Phosphoenolpyruvate carboxylase in rat liver at birth. Biochem J. 1972 Mar;126(5):1127–1134. doi: 10.1042/bj1261127. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Punekar N. S., Lardy H. A. Phosphoenolpyruvate carboxykinase ferroactivator 1. Mechanism of action and identity with glutathione peroxidase. J Biol Chem. 1987 May 15;262(14):6714–6719. [PubMed] [Google Scholar]
  29. Riou J. P., Claus T. H., Pilkis S. J. Stimulation of glucagon of in vivo phosphorylation of rat hepatic pyruvate kinase. J Biol Chem. 1978 Feb 10;253(3):656–659. [PubMed] [Google Scholar]
  30. Rognstad R., Katz J. Role of pyruvate kinase in the regulation of gluconeogenesis from L-lactate. J Biol Chem. 1977 Mar 25;252(6):1831–1833. [PubMed] [Google Scholar]
  31. Schramm V. L., Fullin F. A., Zimmerman M. D. Kinetic studies of the interaction of substrates, Mn2+, and Mg2+ with the Mn2+-sensitive and -insensitive forms of phosphoenolpyruvate carboxykinase. J Biol Chem. 1981 Nov 10;256(21):10803–10808. [PubMed] [Google Scholar]
  32. Scrutton M. C., White M. D. Pyruvate carboxylase. Inhibition of the mammalian and avian liver enzymes by alpha-ketoglutarate and L-glutamate. J Biol Chem. 1974 Sep 10;249(17):5405–5415. [PubMed] [Google Scholar]
  33. Siess E. A., Brocks D. G., Lattke H. K., Wieland O. H. Effect of glucagon on metabolite compartmentation in isolated rat liver cells during gluconeogenesis from lactate. Biochem J. 1977 Aug 15;166(2):225–235. doi: 10.1042/bj1660225. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Sistare F. D., Haynes R. C., Jr Estimation of the relative contributions of enhanced production of oxalacetate and inhibition of pyruvate kinase to acute hormonal stimulation of gluconeogenesis in rat hepatocytes. An analysis of the effects of glucagon, angiotensin II, and dexamethasone on gluconeogenic flux from lactate/pyruvate. J Biol Chem. 1985 Oct 15;260(23):12761–12768. [PubMed] [Google Scholar]
  35. Snoke R. E., Johnston J. B., Lardy H. A. Response of phosphopyruvate carboxylase to tryptophan metabolites and metal ions. Eur J Biochem. 1971 Dec;24(2):342–346. doi: 10.1111/j.1432-1033.1971.tb19692.x. [DOI] [PubMed] [Google Scholar]
  36. Staddon J. M., McGivan J. D. Distinct effects of glucagon and vasopressin on proline metabolism in isolated hepatocytes. The role of oxoglutarate dehydrogenase. Biochem J. 1984 Jan 15;217(2):477–483. doi: 10.1042/bj2170477. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Strzelecki T., Strzelecka D., Koch C. D., LaNoue K. F. Sites of action of glucagon and other Ca2+ mobilizing hormones on the malate aspartate cycle. Arch Biochem Biophys. 1988 Jul;264(1):310–320. doi: 10.1016/0003-9861(88)90599-1. [DOI] [PubMed] [Google Scholar]
  38. Ui M., Claus T. H., Exton J. H., Park C. R. Studies on the mechanism of action of glucagon on gluconeogenesis. J Biol Chem. 1973 Aug 10;248(15):5344–5349. [PubMed] [Google Scholar]
  39. Ui M., Exton J. H., Park C. R. Effects of glucagon on glutamate metabolism in the perfused rat liver. J Biol Chem. 1973 Aug 10;248(15):5350–5359. [PubMed] [Google Scholar]
  40. Wicks W. D., Lewis W., McKibbin J. B. Induction of phosphoenolpyruvate carboxykinase by N 6 , O 2 '-dibutyryl cyclic AMP in rat liver. Biochim Biophys Acta. 1972 Mar 30;264(1):177–185. doi: 10.1016/0304-4165(72)90129-8. [DOI] [PubMed] [Google Scholar]
  41. Williamson J. R., Browning E. T., Thurman R. G., Scholz R. Inhibition of glucagon effects in perfused rat liver by (+)decanoylcarnitine. J Biol Chem. 1969 Sep 25;244(18):5055–5064. [PubMed] [Google Scholar]
  42. YOSHIDA A. ENZYMIC PROPERTIES OF MALATE DEHYDROGENASE OF BACILLUS SUBTILIS. J Biol Chem. 1965 Mar;240:1118–1124. [PubMed] [Google Scholar]

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