The enzymic activity of the mammalian pyruvate dehydrogenase complex is regulated by the phosphorylation of three serine residues (sites 1, 2 and 3) located on the E1 component of the complex. Here we report that the four isoenzymes of protein kinase responsible for the phosphorylation and inactivation of pyruvate dehydrogenase (PDK1, PDK2, PDK3 and PDK4) differ in their abilities to phosphorylate the enzyme. PDK1 can phosphorylate all three sites, whereas PDK2, PDK3 and PDK4 each phosphorylate only site 1 and site 2. Although PDK2 phosphorylates site 1 and 2, it incorporates less phosphate in site 2 than PDK3 or PDK4. As a result, the amount of phosphate incorporated by each isoenzyme decreases in the order PDK1>PDK3PDK4>PDK2. Significantly, binding of the coenzyme thiamin pyrophosphate to pyruvate dehydrogenase alters the rates and stoichiometries of phosphorylation of the individual sites. First, the rate of phosphorylation of site 1 by all isoenzymes of kinase is decreased. Secondly, thiamin pyrophosphate markedly decreases the amount of phosphate that PDK1 incorporates in sites 2 and 3 and that PDK2 incorporates in site 2. In contrast, the coenzyme does not significantly affect the total amount of phosphate incorporated in site 2 by PDK3 and PDK4, but instead decreases the rate of phosphorylation of this site. Furthermore, pyruvate dehydrogenase complex phosphorylated by the individual isoenzymes of kinase is reactivated at different rates by pyruvate dehydrogenase phosphatase. Both isoenzymes of phosphatase (PDP1 and PDP2) readily reactivate the complex phosphorylated by PDK2. When pyruvate dehydrogenase is phosphorylated by other isoenzymes, the rates of reactivation decrease in the order PDK4PDK3> PDK1. Taken together, results reported here strongly suggest that the major determinants of the activity state of pyruvate dehydrogenase in mammalian tissues include the phosphorylation site specificity of isoenzymes of kinase in addition to the absolute amounts of kinase and phosphatase protein expressed in mitochondria.
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August 2001
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Research Article|
August 08 2001
Regulation of pyruvate dehydrogenase activity through phosphorylation at multiple sites
Elena KOLOBOVA;
Elena KOLOBOVA
Division of Molecular Biology and Biochemistry, School of Biological Sciences, University of Missouri–Kansas City, Kansas City, MO 64110-2499, U.S.A.
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Alina TUGANOVA;
Alina TUGANOVA
1
Division of Molecular Biology and Biochemistry, School of Biological Sciences, University of Missouri–Kansas City, Kansas City, MO 64110-2499, U.S.A.
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Igor BOULATNIKOV;
Igor BOULATNIKOV
Division of Molecular Biology and Biochemistry, School of Biological Sciences, University of Missouri–Kansas City, Kansas City, MO 64110-2499, U.S.A.
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Kirill M. POPOV
Kirill M. POPOV
2
Division of Molecular Biology and Biochemistry, School of Biological Sciences, University of Missouri–Kansas City, Kansas City, MO 64110-2499, U.S.A.
2To whom correspondence should be addressed (e-mail popovk@umkc.edu).
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Biochem J (2001) 358 (1): 69–77.
Article history
Received:
February 07 2001
Revision Received:
April 20 2001
Accepted:
June 08 2001
Citation
Elena KOLOBOVA, Alina TUGANOVA, Igor BOULATNIKOV, Kirill M. POPOV; Regulation of pyruvate dehydrogenase activity through phosphorylation at multiple sites. Biochem J 15 August 2001; 358 (1): 69–77. doi: https://doi.org/10.1042/bj3580069
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