Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 1986 Oct 15;239(2):347–354. doi: 10.1042/bj2390347

Kinase activator protein mediates longer-term effects of starvation on activity of pyruvate dehydrogenase kinase in rat liver mitochondria.

G S Denyer, A L Kerbey, P J Randle
PMCID: PMC1147287  PMID: 3814076

Abstract

Starvation of rats for 48 h increased the activity of PDH (pyruvate dehydrogenase) kinase 2.2-fold in extracts of liver mitochondria, 2.9-fold in PDH complex partially purified therefrom by fractional precipitation, and 5-fold in PDH complex partially purified by gel filtration on Sephacryl S-300. A protein fraction was separated from PDH complex in extracts of rat liver mitochondria by gel filtration or fractional precipitation, which increased the activity of PDH kinase in rat liver and pig heart PDH complexes. The activity of this protein fraction was increased approx. 2.5-fold by 48 h starvation of rats. With highly purified pig heart PDH complex it was shown that the protein fraction increased the Vmax. of the PDH kinase reaction 35-fold (fraction from fed rats) or 82-fold (fraction from starved rats); starvation had no effect on the concentration of protein fraction required to give 0.5 Vmax. Evidence is given that the increase in PDH kinase activity effected in extracts of liver mitochondria by starvation is due to increased activity of kinase activator protein, which is tightly bound by rat liver PDH complex and not removed by a single gel filtration. With pig heart PDH complex, increased PDH kinase activity was retained after gel filtration of an admixture with kinase activator protein from starved rats, but was restored to the control value by a second gel filtration; the alterations in PDH kinase activity were associated with obvious changes in protein bands in SDS gels.

Full text

PDF
347

Images in this article

352Fig. 3.
on p.352
353Fig. 4.
on p.353

Selected References

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

  1. Agius L., Alberti K. G. Regulation of flux through pyruvate dehydrogenase and pyruvate carboxylase in rat hepatocytes. Effects of fatty acids and glucagon. Eur J Biochem. 1985 Nov 4;152(3):699–707. doi: 10.1111/j.1432-1033.1985.tb09250.x. [DOI] [PubMed] [Google Scholar]
  2. Baxter M. A., Coore H. G. The mode of regulation of pyruvate dehydrogenase of lactating rat mammary gland. Effects of starvation and insulin. Biochem J. 1978 Aug 15;174(2):553–561. doi: 10.1042/bj1740553. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  4. Caterson I. D., Fuller S. J., Randle P. J. Effect of the fatty acid oxidation inhibitor 2-tetradecylglycidic acid on pyruvate dehydrogenase complex activity in starved and alloxan-diabetic rats. Biochem J. 1982 Oct 15;208(1):53–60. doi: 10.1042/bj2080053. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Cooper R. H., Randle P. J., Denton R. M. Regulation of heart muscle pyruvate dehydrogenase kinase. Biochem J. 1974 Dec;143(3):625–641. doi: 10.1042/bj1430625. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Corbin J. D., Reimann E. M. Assay of cyclic AMP-dependent protein kinases. Methods Enzymol. 1974;38:287–290. doi: 10.1016/0076-6879(74)38044-5. [DOI] [PubMed] [Google Scholar]
  7. De Marcucci O. L., Hunter A., Lindsay J. G. Low immunogenicity of the common lipoamide dehydrogenase subunit (E3) of mammalian pyruvate dehydrogenase and 2-oxoglutarate dehydrogenase multienzyme complexes. Biochem J. 1985 Mar 1;226(2):509–517. doi: 10.1042/bj2260509. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Dennis S. C., DeBuysere M., Scholz R., Olson M. S. Studies on the relationship between ketogenesis and pyruvate oxidation in isolated rat liver mitochondria. J Biol Chem. 1978 Apr 10;253(7):2229–2237. [PubMed] [Google Scholar]
  9. Denton R. M., Randle P. J., Martin B. R. Stimulation by calcium ions of pyruvate dehydrogenase phosphate phosphatase. Biochem J. 1972 Jun;128(1):161–163. doi: 10.1042/bj1280161. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Espinal J., Beggs M., Patel H., Randle P. J. Effects of low-protein diet and starvation on the activity of branched-chain 2-oxo acid dehydrogenase kinase in rat liver and heart. Biochem J. 1986 Jul 1;237(1):285–288. doi: 10.1042/bj2370285. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Fatania H. R., Vary T. C., Randle P. J. Modulation of pyruvate dehydrogenase kinase activity in cultured hepatocytes by glucagon and n-octanoate. Biochem J. 1986 Feb 15;234(1):233–236. doi: 10.1042/bj2340233. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Fuller S. J., Randle P. J. Reversible phosphorylation of pyruvate dehydrogenase in rat skeletal-muscle mitochondria. Effects of starvation and diabetes. Biochem J. 1984 Apr 15;219(2):635–646. doi: 10.1042/bj2190635. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Hutson N. J., Randle P. J. Enhanced activity of pyruvate dehydrogenase kinase in rat heart mitochondria in alloxan-diabetes or starvation. FEBS Lett. 1978 Aug 1;92(1):73–76. doi: 10.1016/0014-5793(78)80724-8. [DOI] [PubMed] [Google Scholar]
  14. Kerbey A. L., Radcliffe P. M., Randle P. J. Diabetes and the control of pyruvate dehydrogenase in rat heart mitochondria by concentration ratios of adenosine triphosphate/adenosine diphosphate, of reduced/oxidized nicotinamide-adenine dinucleotide and of acetyl-coenzyme A/coenzyme A. Biochem J. 1977 Jun 15;164(3):509–519. doi: 10.1042/bj1640509. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kerbey A. L., Radcliffe P. M., Randle P. J., Sugden P. H. Regulation of kinase reactions in pig heart pyruvate dehydrogenase complex. Biochem J. 1979 Aug 1;181(2):427–433. doi: 10.1042/bj1810427. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kerbey A. L., Randle P. J. Pyruvate dehydrogenase kinase activity of pig heart pyruvate dehydrogenase (E1 component of pyruvate dehydrogenase complex). Biochem J. 1985 Nov 1;231(3):523–529. doi: 10.1042/bj2310523. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Kerbey A. L., Randle P. J. Pyruvate dehydrogenase kinase/activator in rat heart mitochondria, Assay, effect of starvation, and effect of protein-synthesis inhibitors of starvation. Biochem J. 1982 Jul 15;206(1):103–111. doi: 10.1042/bj2060103. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kerbey A. L., Randle P. J. Thermolabile factor accelerates pyruvate dehydrogenase kinase reaction in heart mitochondria of starved or alloxan-diabetic rats. FEBS Lett. 1981 May 18;127(2):188–192. doi: 10.1016/0014-5793(81)80201-3. [DOI] [PubMed] [Google Scholar]
  19. Kerbey A. L., Richardson L. J., Randle P. J. The roles of intrinsic kinase and of kinase/activator protein in the enhanced phosphorylation of pyruvate dehydrogenase complex in starvation. FEBS Lett. 1984 Oct 15;176(1):115–119. doi: 10.1016/0014-5793(84)80923-0. [DOI] [PubMed] [Google Scholar]
  20. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  21. Linn T. C., Pettit F. H., Hucho F., Reed L. J. Alpha-keto acid dehydrogenase complexes. XI. Comparative studies of regulatory properties of the pyruvate dehydrogenase complexes from kidney, heart, and liver mitochondria. Proc Natl Acad Sci U S A. 1969 Sep;64(1):227–234. doi: 10.1073/pnas.64.1.227. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Linn T. C., Pettit F. H., Reed L. J. Alpha-keto acid dehydrogenase complexes. X. Regulation of the activity of the pyruvate dehydrogenase complex from beef kidney mitochondria by phosphorylation and dephosphorylation. Proc Natl Acad Sci U S A. 1969 Jan;62(1):234–241. doi: 10.1073/pnas.62.1.234. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. 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]
  24. Patston P. A., Espinal J., Randle P. J. Effects of diet and of alloxan-diabetes on the activity of branched-chain 2-oxo acid dehydrogenase complex and of activator protein in rat tissues. Biochem J. 1984 Sep 15;222(3):711–719. doi: 10.1042/bj2220711. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Rahmatullah M., Roche T. E. Modification of bovine kidney pyruvate dehydrogenase kinase activity by CoA esters and their mechanism of action. J Biol Chem. 1985 Aug 25;260(18):10146–10152. [PubMed] [Google Scholar]
  26. Randle P. J. Fuel selection in animals. Biochem Soc Trans. 1986 Oct;14(5):799–806. doi: 10.1042/bst0140799. [DOI] [PubMed] [Google Scholar]
  27. Sale G. J., Randle P. J. Analysis of site occupancies in [32P]phosphorylated pyruvate dehydrogenase complexes by aspartyl-prolyl cleavage of tryptic phosphopeptides. Eur J Biochem. 1981 Dec;120(3):535–540. doi: 10.1111/j.1432-1033.1981.tb05733.x. [DOI] [PubMed] [Google Scholar]
  28. Sale G. J., Randle P. J. Occupancy of phosphorylation sites in pyruvate dehydrogenase phosphate complex in rat heart in vivo. Relation to proportion of inactive complex and rate of re-activation by phosphatase. Biochem J. 1982 Aug 15;206(2):221–229. doi: 10.1042/bj2060221. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Sale G. J., Randle P. J. Role of individual phosphorylation sites in inactivation of pyruvate dehydrogenase complex in rat heart mitochondria. Biochem J. 1982 Apr 1;203(1):99–108. doi: 10.1042/bj2030099. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Scholz R., Olson M. S., Schwab A. J., Schwabe U., Noell C., Braun W. The effect of fatty acids on the regulation of pyruvate dehydrogenase in perfused rat liver. Eur J Biochem. 1978 May 16;86(2):519–530. doi: 10.1111/j.1432-1033.1978.tb12335.x. [DOI] [PubMed] [Google Scholar]
  31. Siess E. A., Wieland O. H. Essential role of coenzyme A in pyruvate dehydrogenase kinase activity. FEBS Lett. 1982 Nov 8;148(2):201–206. doi: 10.1016/0014-5793(82)80808-9. [DOI] [PubMed] [Google Scholar]
  32. Stepp L. R., Pettit F. H., Yeaman S. J., Reed L. J. Purification and properties of pyruvate dehydrogenase kinase from bovine kidney. J Biol Chem. 1983 Aug 10;258(15):9454–9458. [PubMed] [Google Scholar]
  33. Sugden P. H., Hutson N. J., Kerbey A. L., Randle P. J. Phosphorylation of additional sites on pyruvate dehydrogenase inhibits its re-activation by pyruvate dehydrogenase phosphate phosphatase. Biochem J. 1978 Feb 1;169(2):433–435. doi: 10.1042/bj1690433. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Sugden P. H., Kerbey A. L., Randle P. J., Waller C. A., Reid K. B. Amino acid sequences around the sites of phosphorylation in the pig heart pyruvate dehydrogenase complex. Biochem J. 1979 Aug 1;181(2):419–426. doi: 10.1042/bj1810419. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Teague W. M., Pettit F. H., Wu T. L., Silberman S. R., Reed L. J. Purification and properties of pyruvate dehydrogenase phosphatase from bovine heart and kidney. Biochemistry. 1982 Oct 26;21(22):5585–5592. doi: 10.1021/bi00265a031. [DOI] [PubMed] [Google Scholar]
  36. WILKINSON G. N. Statistical estimations in enzyme kinetics. Biochem J. 1961 Aug;80:324–332. doi: 10.1042/bj0800324. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Wieland O. H., Siess E. A., Weiss L., Löffler G., Patzelt C., Portenhauser R., Hartmann U., Schirmann A. Regulation of the mammalian pyruvate dehydrogenase complex by covalent modification. Symp Soc Exp Biol. 1973;27:371–400. [PubMed] [Google Scholar]
  38. Wieland O. H. The mammalian pyruvate dehydrogenase complex: structure and regulation. Rev Physiol Biochem Pharmacol. 1983;96:123–170. doi: 10.1007/BFb0031008. [DOI] [PubMed] [Google Scholar]
  39. Yeaman S. J., Hutcheson E. T., Roche T. E., Pettit F. H., Brown J. R., Reed L. J., Watson D. C., Dixon G. H. Sites of phosphorylation on pyruvate dehydrogenase from bovine kidney and heart. Biochemistry. 1978 Jun 13;17(12):2364–2370. doi: 10.1021/bi00605a017. [DOI] [PubMed] [Google Scholar]
  40. Zwiebel F. M., Schwabe U., Olson M. S., Scholz R. Role of pyruvate transporter in the regulation of the pyruvate dehydrogenase multienzyme complex in perfused rat liver. Biochemistry. 1982 Jan 19;21(2):346–353. doi: 10.1021/bi00531a023. [DOI] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES