Bovine heart pyruvate dehydrogenase kinase stimulation by alpha-ketoisovalerate.
Purified bovine heart pyruvate dehydrogenase complex was used to investigate the effects of monovalent cations and alpha-ketoisovalerate on pyruvate dehydrogenase (PDH) kinase inhibition by thiamin pyrophosphate. Initial velocity patterns for thiamin pyrophosphate inhibition were consistent with hyperbolic non-competitive or hyperbolic uncompetitive inhibition at various K+ concentrations between 0 and 120 mM. The Kis, Kid, and Kin for thiamin pyrophosphate were in the range of 0.009 to 5.1 microM over the range of K+ concentrations tested. In the absence of K+, 1 mM alpha-ketoisovalerate had no effect on PDH kinase inhibition by thiamin pyrophosphate, whereas in the presence of 20 mM K+, alpha-ketoisovalerate stimulated PDH kinase activity almost 2-fold over the range of 0-80 microM thiamin pyrophosphate. Half-maximal stimulation by alpha-ketoisovalerate occurred at about 200 microM in the presence of 100 microM thiamin pyrophosphate and 20 mM K+. Similar but less extensive changes occurred in the presence of 100 microM thiamin pyrophosphate and 1 mM NH4+. Initial velocity patterns for PDH kinase inhibition by thiamin pyrophosphate in the presence of 2 mM alpha-ketoisovalerate were mixed noncompetitive, but alpha-ketoisovalerate increased the Vm and Km for adenosine 5'-triphosphate in the presence of inhibitor. In the presence of thiamin pyrophosphate, PDH kinase remained stimulated after chromatography on Sephadex G-25 to remove alpha-ketoisovalerate. The results indicate that acylation of pyruvate dehydrogenase complex by alpha-ketoisovalerate results in PDH kinase stimulation but only in the presence of monovalent cations and thiamin pyrophosphate.
Department of Biochemistry, University of Texas Health Science Center, San Antonio 78284.
J G
JG
Robertson
L L
LL
Barron
M S
MS
Olson
D000215:Acylation; D000818:Animals; D002414:Cations, Monovalent; D002417:Cattle; D045782:Hemiterpenes; D007651:Keto Acids / Q000494:pharmacology*; D007700:Kinetics; D008433:Mathematics; D008954:Models, Biological; D009206:Myocardium / Q000201:enzymology*; D011189:Potassium Chloride / Q000494:pharmacology; D011494:Protein Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D000081382:Pyruvate Dehydrogenase Acetyl-Transferring Kinase; D013835:Thiamine Pyrophosphate / Q000494:pharmacology
D002414:Cations, Monovalent; D045782:Hemiterpenes; D007651:Keto Acids; D000081382:Pyruvate Dehydrogenase Acetyl-Transferring Kinase; D011189:Potassium Chloride; C001505:alpha-ketoisovalerate; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D013835:Thiamine Pyrophosphate