Increased hepatic pyruvate dehydrogenase kinase activity in fed hyperthyroid rats: studies in vivo and with cultured hepatocytes

1996 ◽  
Vol 119 (2) ◽  
pp. 219-224 ◽  
Author(s):  
Mary C. Sugden ◽  
Lee G.D. Fryer ◽  
David A. Priestman ◽  
Karen A. Orfali ◽  
Mark J. Holness
Keyword(s):  
Pyruvate Dehydrogenase ◽  
Kinase Activity ◽  
Pyruvate Dehydrogenase Kinase ◽  
Cultured Hepatocytes

scholarly journals Regulation of pyruvate dehydrogenase kinase activity from pig kidney cortex

1992 ◽  
Vol 288 (2) ◽  
pp. 369-373 ◽  
Author(s):  
T Pawelczyk ◽  
M S Olson
Keyword(s):  
Ionic Strength ◽  
Pyruvate Dehydrogenase ◽  
Kinase Activity ◽  
Pyruvate Dehydrogenase Kinase ◽  
Kidney Cortex ◽  
Small Range ◽  
High Concentration ◽  
Optimum Activity ◽  
Pig Kidney

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.

scholarly journals Insulin reverses effects of starvation on the activity of pyruvate dehydrogenase kinase in cultured hepatocytes

1987 ◽  
Vol 246 (1) ◽  
pp. 233-236 ◽  
Author(s):  
D R Marchington ◽  
A L Kerbey ◽  
A E Jones ◽  
P J Randle
Keyword(s):  
Tissue Culture ◽  
Pyruvate Dehydrogenase ◽  
Pyruvate Dehydrogenase Kinase ◽  
Cultured Hepatocytes ◽  
Activator Protein ◽  
Culture Of Hepatocytes

In tissue culture of hepatocytes, insulin (0.1-1 munits/ml for 4 h) reversed completely the effects of starvation of rats to decrease the activity of pyruvate dehydrogenase (PDH) complex and to increase the activities of PDH kinase and PDH kinase activator protein. It had no effect in hepatocytes from fed rats. Significant effects of insulin were detected with 0.01 munit/ml after 4 h, and in 1-2 h with 1 munit/ml.

scholarly journals Studies of the long-term regulation of hepatic pyruvate dehydrogenase kinase

1998 ◽  
Vol 329 (1) ◽  
pp. 89-94 ◽  
Author(s):  
C. Mary SUGDEN ◽  
G. D. Lee FRYER ◽  
A. Karen ORFALI ◽  
A. David PRIESTMAN ◽  
Elaine DONALD ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Pyruvate Dehydrogenase ◽  
Unsaturated Fatty Acids ◽  
Fold Increase ◽  
Membrane Lipid ◽  
Dietary Lipid ◽  
Pyruvate Dehydrogenase Kinase ◽  
Plasma Insulin Concentration

The administration of a low-carbohydrate/high-saturated-fat (LC/HF) diet for 28 days or starvation for 48 h both increased pyruvate dehydrogenase kinase (PDHK) activity in extracts of rat hepatic mitochondria, by approx. 2.1-fold and 3.5-fold respectively. ELISAs of extracts of hepatic mitochondria, conducted over a range of pyruvate dehydrogenase (PDH) activities, revealed that mitochondrial immunoreactive PDHKII (the major PDHK isoform in rat liver) was significantly increased by approx. 1.4-fold after 28 days of LC/HF feeding and by approx. 2-fold after 48 h of starvation. The effect of LC/HF feeding to increase hepatic PDHK activity was retained through hepatocyte preparation, but was decreased on 21 h culture with insulin (100μ-i.u./ml). A sustained (24 h) 2-4-fold elevation in plasma insulin concentration in vivo (achieved by insulin infusion via an osmotic pump) suppressed the effect of LC/HF feeding so that hepatic PDHK activities did not differ significantly from those of (insulin-infused) control rats. The increase in hepatic PDHK activity evoked by 28 days of LC/HF feeding was prevented and reversed (within 24 h) by the replacement of 7% of the dietary lipid with long-chain ω-3 fatty acids. Analysis of hepatic membrane lipid revealed a 1.9-fold increase in the ratio of total polyunsaturated ω-3 fatty acids to total mono-unsaturated fatty acids. The results indicate that the increased hepatic PDHK activities observed in livers of LC/HF-fed or 48 h-starved rats are associated with long-term actions to increase hepatic PDHKII concentrations. The long-term regulation of hepatic PDHK by LC/HF feeding might be achieved through an impaired action of insulin to suppress PDHK activity. In addition, the fatty acid composition of the diet, rather than the fat content, is a key influence.

Increased pyruvate dehydrogenase kinase activity in response to sepsis

1991 ◽  
Vol 260 (5) ◽  
pp. E669-E674 ◽  
Author(s):  
T. C. Vary
Keyword(s):  
Skeletal Muscle ◽  
Pyruvate Dehydrogenase ◽  
Kinase Activity ◽  
Pyruvate Dehydrogenase Complex ◽  
Pyruvate Dehydrogenase Kinase ◽  
Sterile Inflammation ◽  
Stable Factor ◽  
Skeletal Muscle Mitochondria ◽  
Dependent Inactivation ◽  
Significant Difference

The effect of sterile inflammation and sepsis on the proportion of active pyruvate dehydrogenase complex (PDH) in mitochondria isolated from skeletal muscle has been investigated. The proportion of active PDH in mitochondria isolated from septic animals was significantly reduced compared with control under all incubation conditions examined, even in the presence of inhibitors of the PDH kinase. There was no significant difference between control and sterile inflammation in any of the incubations examined. The rate constant for ATP-dependent inactivation of the PDH complex in mitochondrial extracts from control animals was -0.42 min-1 (r = 0.993; P less than 0.001) and was not altered in mitochondrial extracts from sterile inflammatory animals (-0.43 min-1; r = 0.999; P less than 0.001). However, rate constants for inactivation in septic animals was significantly increased over twofold to -1.08 min-1 (r = 0.987; P less than 0.001) (P less than 0.001 vs. control or sterile inflammation). In the presence of inhibitors of the PDH kinase reaction (2.5 mM pyruvate or 1 mM dichloroacetate), inactivation of PDH after addition of ATP was significantly greater in mitochondrial extracts from septic than either control or sterile inflammatory animals. These results suggest that sepsis, but not sterile inflammation, induces a stable factor in skeletal muscle mitochondria that increased PDH kinase activity.

scholarly journals Phosphorylation state of cytosolic and mitochondrial adenine nucleotides and of pyruvate dehydrogenase in isolated rat liver cells

1976 ◽  
Vol 156 (1) ◽  
pp. 91-102 ◽  
Author(s):  
E A Siess ◽  
O H Wieland
Keyword(s):  
Pyruvate Dehydrogenase ◽  
Adenine Nucleotides ◽  
Active Form ◽  
Inverse Correlation ◽  
Liver Cells ◽  
Diabetic Rats ◽  
Pyruvate Dehydrogenase Kinase ◽  
Intact Cells ◽  
Phosphorylation State

1. Cytosolic and mitochondrial ATP and ADP concentrations of liver cells isolated from normal fed, starved and diabetic rats were determined. 2. The cytosolic ATP/ADP ratio was 6,9 and 10 in normal fed, starved and diabetic rats respectively. 3. The mitochondrial ATP/ADP ratio was 2 in normal and diabetic rats and 1.6 in starved rats. 4. Adenosine increased the cytosolic and lowered the mitochondrial ATP/ADP ratio, whereas atractyloside had the opposite effect. 5. Incubation of the hepatocytes with fructose, glycerol or sorbitol led to a fall in the ATP/ADP ratio in both the cytosolic and the mitochondrial compartment. 6. The interrelationship between the mitochondrial ATP/ADP ratio and the phosphorylation state of pyruvate dehydrogenase in intact cells was studied. 7. In hepatocytes isolated from fed rats an inverse correlation between the mitochondrial ATP/ADP ratio and the active form of pyruvate dehydrogenase (pyruvate dehydrogenase a) was demonstrable on loading with fructose, glycerol or sorbitol. 8. No such correlation was obtained with pyruvate or dihydroxyacetone. For pyruvate, this can be explained by inhibition of pyruvate dehydrogenase kinase. 9. Liver cells isolated from fed animals displayed pyruvate dehydrogenase a activity twice that found in vivo. Physiological values were obtained when the hepatocytes were incubated with albumin-oleate, which also yielded the highest mitochondrial ATP/ADP ratio.

scholarly journals Interaction between the individual isoenzymes of pyruvate dehydrogenase kinase and the inner lipoyl-bearing domain of transacetylase component of pyruvate dehydrogenase complex

2002 ◽  
Vol 366 (1) ◽  
pp. 129-136 ◽  
Author(s):  
Alina TUGANOVA ◽  
Igor BOULATNIKOV ◽  
Kirill M. POPOV
Keyword(s):  
Pyruvate Dehydrogenase ◽  
Kinase Activity ◽  
Pyruvate Dehydrogenase Complex ◽  
Tight Binding ◽  
Enzymic Activity ◽  
Pyruvate Dehydrogenase Kinase ◽  
Size Exclusion ◽  
Apparent Dissociation Constant ◽  
The Individual ◽  
Dehydrogenase Complex

Protein—protein interactions play an important role in the regulation of enzymic activity of pyruvate dehydrogenase kinase (PDK). It is generally believed that the binding of PDK to the inner lipoyl-bearing domain L2 of the transacetylase component E2 of pyruvate dehydrogenase complex largely determines the level of kinase activity. In the present study, we characterized the interaction between the individual isoenzymes of PDK (PDK1—PDK4) and monomeric L2 domain of human E2, as well as the effect of this interaction on kinase activity. It was found that PDK isoenzymes are markedly different with respect to their affinities for L2. PDK3 demonstrated a very tight binding, which persisted during isolation of PDK3—L2 complexes using size-exclusion chromatography. Binding of PDK1 and PDK2 was readily reversible with the apparent dissociation constant of approx. 10μM for both isoenzymes. PDK4 had a greatly reduced capacity for L2 binding (relative order PDK3>PDK1 = PDK2>PDK4). Monomeric L2 domain alone had very little effect on the activities of either PDK1 or PDK2. In contrast, L2 caused a 3-fold increase in PDK3 activity and approx. 37% increase in PDK4 activity. These results strongly suggest that the interactions between the individual isoenzymes of PDK and L2 domain are isoenzyme-specific and might be among the major factors that determine the level of kinase activity of particular isoenzyme towards the pyruvate dehydrogenase complex.

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