Regulation of pyruvate dehydrogenase kinase activity from pig kidney cortex.
The activity of pyruvate dehydrogenase (PDH) kinase in the purified PDH complex from pig kidney is sensitive to changes in ionic strength. The enzyme has optimum activity within a small range of ionic strength (0.03-0.05 M). An increase in ionic strength from 0.04 M to 0.2 M lowers the activity of PDH kinase by 32% and decreases the Km for ATP from 25 microM to 10 microM. At constant ionic strength (0.15 M) the enzyme has optimum activity over a broad pH range (7.2-8.0). The PDH kinase is stimulated 2.2-fold by 20 mM-K+, whereas Na+ even at high concentration (80 mM) has no effect on the enzyme activity. The stimulation of PDH kinase by K+ is not dependent on pH and ionic strength. PDH kinase is inhibited by HPO4(2-) in the presence of K+, whereas HPO4(2-) has no effect on the activity of this enzyme in the absence of K+. HPO4(2-) at concentrations of 2 and 10 mM inhibits PDH kinase by 28% and 55% respectively. The magnitude of this inhibition is not dependent on the ATP/ADP ratio. Inhibition by HPO4(2-) in the concentration range 0-10 mM is non-competitive with respect to ATP, and becomes mixed-type at concentrations over 10 mM. The Ki for HPO4(2-) is 10 mM. When HPO4(2-) is replaced by SO4(2-), the same effects on the activity of PDH kinase are observed. PDH kinase is also inhibited by Cl-. In the presence of 80 mM-Cl- the PDH kinase is inhibited by 40%. The inhibition by Cl- is not dependent on K+. In conclusion, we postulate that changes in phosphate concentrations may play a significant role in the regulation of PDH kinase activity in vivo.
Pawelczyk
T
T
Department of Biochemistry, University of Texas Health Science Center, San Antonio 78284-7760.
Olson
M S
MS
D000818:Animals; D002712:Chlorides / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007672:Kidney Cortex / Q000201:enzymology*; D007700:Kinetics; D008274:Magnesium / Q000378:metabolism; D009994:Osmolar Concentration; D010710:Phosphates / Q000378:metabolism*; D011188:Potassium / Q000378:metabolism; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D000081382:Pyruvate Dehydrogenase Acetyl-Transferring Kinase; D011768:Pyruvate Dehydrogenase Complex / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism; D013431:Sulfates / Q000378:metabolism; D013552:Swine
D002712:Chlorides; D010710:Phosphates; D047428:Protein Kinase Inhibitors; D000081382:Pyruvate Dehydrogenase Acetyl-Transferring Kinase; D011768:Pyruvate Dehydrogenase Complex; D013431:Sulfates; D012964:Sodium; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D008274:Magnesium; D011188:Potassium