scholarly journals Identification of a novel malonyl-CoA IC50 for CPT-I: implications for predicting in vivo fatty acid oxidation rates

2012 ◽  
Vol 448 (1) ◽  
pp. 13-20 ◽  
Author(s):  
Brennan K. Smith ◽  
Christopher G. R. Perry ◽  
Timothy R. Koves ◽  
David C. Wright ◽  
Jeffrey C. Smith ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Fatty Acid Oxidation ◽  
Acid Oxidation ◽  
Muscle Fibres ◽  
Inhibition Kinetics ◽  
Oxidation Rates ◽  
Isolated Mitochondria ◽  
Malonyl Coa ◽  
Cpt I

Published values regarding the sensitivity (IC50) of CPT-I (carnitine palmitoyltransferase I) to M-CoA (malonyl-CoA) inhibition in isolated mitochondria are inconsistent with predicted in vivo rates of fatty acid oxidation. Therefore we have re-examined M-CoA inhibition kinetics under various P-CoA (palmitoyl-CoA) concentrations in both isolated mitochondria and PMFs (permeabilized muscle fibres). PMFs have an 18-fold higher IC50 (0.61 compared with 0.034 μM) in the presence of 25 μM P-CoA and a 13-fold higher IC50 (6.3 compared with 0.49 μM) in the presence of 150 μM P-CoA compared with isolated mitochondria. M-CoA inhibition kinetics determined in PMFs predicts that CPT-I activity is inhibited by 33% in resting muscle compared with >95% in isolated mitochondria. Additionally, the ability of M-CoA to inhibit CPT-I appears to be dependent on P-CoA concentration, as the relative inhibitory capacity of M-CoA is decreased with increasing P-CoA concentrations. Altogether, the use of PMFs appears to provide an M-CoA IC50 that better reflects the predicted in vivo rates of fatty acid oxidation. These findings also demonstrate that the ratio of [P-CoA]/[M-CoA] is critical for regulating CPT-I activity and may partially rectify the in vivo disconnect between M-CoA content and CPT-I flux within the context of exercise and Type 2 diabetes.

Effect of phosphorylation by AMP-activated protein kinase on palmitoyl-CoA inhibition of skeletal muscle acetyl-CoA carboxylase

2005 ◽  
Vol 98 (4) ◽  
pp. 1221-1227 ◽  
Author(s):  
D. S. Rubink ◽  
W. W. Winder
Keyword(s):  
Skeletal Muscle ◽  
Fatty Acid ◽  
Protein Kinase ◽  
Fatty Acid Oxidation ◽  
Acid Oxidation ◽  
Acetyl Coa Carboxylase ◽  
Acetyl Coa ◽  
Oxidation Rates ◽  
Malonyl Coa ◽  
Amp Activated Protein Kinase

AMP-activated protein kinase (AMPK) has previously been demonstrated to phosphorylate and inactivate skeletal muscle acetyl-CoA carboxylase (ACC), the enzyme responsible for synthesis of malonyl-CoA, an inhibitor of carnitine palmitoyltransferase 1 and fatty acid oxidation. Contraction-induced activation of AMPK with subsequent phosphorylation/inactivation of ACC has been postulated to be responsible in part for the increase in fatty acid oxidation that occurs in muscle during exercise. These studies were designed to answer the question: Does phosphorylation of ACC by AMPK make palmitoyl-CoA a more effective inhibitor of ACC? Purified rat muscle ACC was subjected to phosphorylation by AMPK. Activity was determined on nonphosphorylated and phosphorylated ACC preparations at acetyl-CoA concentrations ranging from 2 to 500 μM and at palmitoyl-CoA concentrations ranging from 0 to 100 μM. Phosphorylation resulted in a significant decline in the substrate saturation curve at all palmitoyl-CoA concentrations. The inhibitor constant for palmitoyl-CoA inhibition of ACC was reduced from 1.7 ± 0.25 to 0.85 ± 0.13 μM as a consequence of phosphorylation. At 0.5 mM citrate, ACC activity was reduced to 13% of control values in response to the combination of phosphorylation and 10 μM palmitoyl-CoA. Skeletal muscle ACC is more potently inhibited by palmitoyl-CoA after having been phosphorylated by AMPK. This may contribute to low-muscle malonyl-CoA values and increasing fatty acid oxidation rates during long-term exercise when plasma fatty acid concentrations are elevated.

Regulation of myocardial fatty acid oxidation by substrate supply

2001 ◽  
Vol 281 (4) ◽  
pp. H1561-H1567 ◽  
Author(s):  
Sarah L. Longnus ◽  
Richard B. Wambolt ◽  
Rick L. Barr ◽  
Gary D. Lopaschuk ◽  
Michael F. Allard
Keyword(s):  
Fatty Acid ◽  
Fatty Acid Oxidation ◽  
Acid Oxidation ◽  
Acetyl Coa ◽  
Regulatory Pathways ◽  
Oxidation Rates ◽  
Substrate Supply ◽  
Exogenous Substrate ◽  
Malonyl Coa ◽  
Myocardial Substrate

We tested the hypothesis that myocardial substrate supply regulates fatty acid oxidation independent of changes in acetyl-CoA carboxylase (ACC) and 5′-AMP-activated protein kinase (AMPK) activities. Fatty acid oxidation was measured in isolated working rat hearts exposed to different concentrations of exogenous long-chain (0.4 or 1.2 mM palmitate) or medium-chain (0.6 or 2.4 mM octanoate) fatty acids. Fatty acid oxidation was increased with increasing exogenous substrate concentration in both palmitate and octanoate groups. Malonyl-CoA content only rose as acetyl-CoA supply from octanoate oxidation increased. The increases in octanoate oxidation and malonyl-CoA content were independent of changes in ACC and AMPK activity, except that ACC activity increased with very high acetyl-CoA supply levels. Our data suggest that myocardial substrate supply is the primary mechanism responsible for alterations in fatty acid oxidation rates under nonstressful conditions and when substrates are present at physiological concentrations. More extreme variations in substrate supply lead to changes in fatty acid oxidation by the additional involvement of intracellular regulatory pathways.

Malonyl-CoA content and fatty acid oxidation in rat muscle and liver in vivo

2000 ◽  
Vol 279 (2) ◽  
pp. E259-E265 ◽  
Author(s):  
David Chien ◽  
David Dean ◽  
Asish K. Saha ◽  
J. P. Flatt ◽  
Neil B. Ruderman
Keyword(s):  
Fatty Acid ◽  
Fatty Acid Oxidation ◽  
Fatty Acyl ◽  
Whole Body ◽  
Acid Oxidation ◽  
Malonyl Coa ◽  
Plasma Ffa ◽  
Gastrocnemius Muscles ◽  
Time Point

Malonyl-CoA acutely regulates fatty acid oxidation in liver in vivo by inhibiting carnitine palmitoyltransferase. Thus rapid increases in the concentration of malonyl-CoA, accompanied by decreases in long-chain fatty acyl carnitine (LCFA-carnitine) and fatty acid oxidation have been observed in liver of fasted-refed rats. It is less clear that it plays a similar role in skeletal muscle. To examine this question, whole body respiratory quotients (RQ) and the concentrations of malonyl-CoA and LCFA-carnitine in muscle were determined in 48-h-starved rats before and at various times after refeeding. RQ values were 0.82 at baseline and increased to 0.93, 1.0, 1.05, and 1.09 after 1, 3, 12, and 18 h of refeeding, respectively, suggesting inhibition of fat oxidation in all tissues. The increases in RQ at each time point correlated closely ( r = 0.98) with increases (50–250%) in the concentration of malonyl-CoA in soleus and gastrocnemius muscles and decreases in plasma FFA and muscle LCFA-carnitine levels. Similar changes in malonyl-CoA and LCFA-carnitine were observed in liver. The increases in malonyl-CoA in muscle during refeeding were not associated with increases in the assayable activity of acetyl-CoA carboxylase (ACC) or decreases in the activity of malonyl-CoA decarboxylase (MCD). The results suggest that, during refeeding after a fast, decreases in fatty acid oxidation occur rapidly in muscle and are attributable both to decreases in plasma FFA and increases in the concentration of malonyl-CoA. They also suggest that the increase in malonyl-CoA in this situation is not due to changes in the assayable activity of either ACC or MCD or an increase in the cytosolic concentration of citrate.

scholarly journals The role of changes in the sensitivity of hepatic mitochondrial overt carnitine palmitoyltransferase in determining the onset of the ketosis of starvation in the rat

1996 ◽  
Vol 318 (3) ◽  
pp. 767-770 ◽  
Author(s):  
Lesley DRYNAN ◽  
Patti A. QUANT ◽  
Victor A. ZAMMIT
Keyword(s):  
Fatty Acid ◽  
Fatty Acid Oxidation ◽  
Time Course ◽  
Ketone Body ◽  
Serum Insulin ◽  
Carnitine Palmitoyltransferase ◽  
Acid Oxidation ◽  
Malonyl Coa ◽  
Cpt I ◽  
Body Concentration

The relationships between the increase in blood ketone-body concentrations and several parameters that can potentially influence the rate of hepatic fatty acid oxidation were studied during progressive starvation (up to 24 h) in the rat in order to discover whether the sensitivity of mitochondrial overt carnitine palmitoyltransferase (CPT I) to malonyl-CoA plays an important part in determining the intrahepatic potential for fatty acid oxidation during the onset of ketogenic conditions. A rapid increase in blood ketone-body concentration occurred between 12 and 16 h of starvation, several hours after the marked fall in hepatic malonyl-CoA and in serum insulin concentrations and doubling of plasma non-esterfied fatty acid (NEFA) concentration. Consequently, both the changes in hepatic malonyl-CoA and serum NEFA preceded the increase in blood ketone-body concentration by several hours. The maximal activity of CPT I increased gradually throughout the 24 h period of starvation, but the increases did not become significant before 18 h of starvation. By contrast, the sensitivity of CPT I to malonyl-CoA and the increase in blood ketone-body concentration followed an identical time course, demonstrating the central importance of this parameter in determining the ketogenic response of the liver to the onset of the starved state.

scholarly journals Evidence that the sensitivity of carnitine palmitoyltransferase I to inhibition by malonyl-CoA is an important site of regulation of hepatic fatty acid oxidation in the fetal and newborn rabbit. Perinatal development and effects of pancreatic hormones in cultured rabbit hepatocytes

1990 ◽  
Vol 269 (2) ◽  
pp. 409-415 ◽  
Author(s):  
C Prip-Buus ◽  
J P Pegorier ◽  
P H Duee ◽  
C Kohl ◽  
J Girard
Keyword(s):  
Fatty Acid ◽  
Cyclic Amp ◽  
Fatty Acid Oxidation ◽  
Carnitine Palmitoyltransferase ◽  
Acid Oxidation ◽  
Fetal Hepatocytes ◽  
Malonyl Coa ◽  
Carnitine Palmitoyltransferase I ◽  
Cpt I ◽  
Hepatic Fatty Acid Oxidation

The temporal changes in oleate oxidation, lipogenesis, malonyl-CoA concentration and sensitivity of carnitine palmitoyltransferase I (CPT 1) to malonyl-CoA inhibition were studied in isolated rabbit hepatocytes and mitochondria as a function of time after birth of the animal or time in culture after exposure to glucagon, cyclic AMP or insulin. (1) Oleate oxidation was very low during the first 6 h after birth, whereas lipogenesis rate and malonyl-CoA concentration decreased rapidly during this period to reach levels as low as those found in 24-h-old newborns that show active oleate oxidation. (2) The changes in the activity of CPT I and the IC50 (concn. causing 50% inhibition) for malonyl-CoA paralleled those of oleate oxidation. (3) In cultured fetal hepatocytes, the addition of glucagon or cyclic AMP reproduced the changes that occur spontaneously after birth. A 12 h exposure to glucagon or cyclic AMP was sufficient to inhibit lipogenesis totally and to cause a decrease in malonyl-CoA concentration, but a 24 h exposure was required to induce oleate oxidation. (4) The induction of oleate oxidation by glucagon or cyclic AMP is triggered by the fall in the malonyl-CoA sensitivity of CPT I. (5) In cultured hepatocytes from 24 h-old newborns, the addition of insulin inhibits no more than 30% of the high oleate oxidation, whereas it stimulates lipogenesis and increases malonyl-CoA concentration by 4-fold more than in fetal cells (no oleate oxidation). This poor effect of insulin on oleate oxidation seems to be due to the inability of the hormone to increase the sensitivity of CPT I sufficiently. Altogether, these results suggest that the malonyl-CoA sensitivity of CPT I is the major site of regulation during the induction of fatty acid oxidation in the fetal rabbit liver.

Subsarcolemmal and intermyofibrillar mitochondria play distinct roles in regulating skeletal muscle fatty acid metabolism

2005 ◽  
Vol 288 (5) ◽  
pp. C1074-C1082 ◽  
Author(s):  
Timothy R. Koves ◽  
Robert C. Noland ◽  
Andrew L. Bates ◽  
Sarah T. Henes ◽  
Deborah M. Muoio ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Fatty Acid ◽  
Exercise Training ◽  
Fatty Acid Oxidation ◽  
Treadmill Running ◽  
Hill Coefficient ◽  
Acid Oxidation ◽  
Oxidation Rates ◽  
Malonyl Coa ◽  
Mitochondrial Fatty Acid

Skeletal muscle contains two populations of mitochondria that appear to be differentially affected by disease and exercise training. It remains unclear how these mitochondrial subpopulations contribute to fiber type-related and/or training-induced changes in fatty acid oxidation and regulation of carnitine palmitoyltransferase-1β (CPT1β), the enzyme that controls mitochondrial fatty acid uptake in skeletal muscle. To this end, we found that fatty acid oxidation rates were 8.9-fold higher in subsarcolemmal mitochondria (SS) and 5.3-fold higher in intermyofibrillar mitochondria (IMF) that were isolated from red gastrocnemius (RG) compared with white gastrocnemius (WG) muscle, respectively. Malonyl-CoA (10 μM), a potent inhibitor of CPT1β, completely abolished fatty acid oxidation in SS and IMF mitochondria from WG, whereas oxidation rates in the corresponding fractions from RG were inhibited only 89% and 60%, respectively. Endurance training also elicited mitochondrial adaptations that resulted in enhanced fatty acid oxidation capacity. Ten weeks of treadmill running differentially increased palmitate oxidation rates 100% and 46% in SS and IMF mitochondria, respectively. In SS mitochondria, elevated fatty acid oxidation rates were accompanied by a 48% increase in citrate synthase activity but no change in CPT1 activity. Nonlinear regression analyses of mitochondrial fatty acid oxidation rates in the presence of 0–100 μM malonyl-CoA indicated that IC50 values were neither dependent on mitochondrial subpopulation nor affected by exercise training. However, in IMF mitochondria, training reduced the Hill coefficient ( P < 0.05), suggesting altered CPT1β kinetics. These results demonstrate that endurance exercise provokes subpopulation-specific changes in mitochondrial function that are characterized by enhanced fatty acid oxidation and modified CPT1β-malonyl-CoA dynamics.

Malonyl-CoA decarboxylase inhibition suppresses fatty acid oxidation and reduces lactate production during demand-induced ischemia

2005 ◽  
Vol 289 (6) ◽  
pp. H2304-H2309 ◽  
Author(s):  
William C. Stanley ◽  
Eric E. Morgan ◽  
Hazel Huang ◽  
Tracy A. McElfresh ◽  
Joseph P. Sterk ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Fatty Acid Oxidation ◽  
Glucose Oxidation ◽  
Contractile Function ◽  
Lactate Production ◽  
Specific Inhibitor ◽  
Acid Oxidation ◽  
Malonyl Coa ◽  
Cpt I

The rate of cardiac fatty acid oxidation is regulated by the activity of carnitine palmitoyltransferase-I (CPT-I), which is inhibited by malonyl-CoA. We tested the hypothesis that the activity of the enzyme responsible for malonyl-CoA degradation, malonyl-CoA decarboxlyase (MCD), regulates myocardial malonyl-CoA content and the rate of fatty acid oxidation during demand-induced ischemia in vivo. The myocardial content of malonyl-CoA was increased in anesthetized pigs using a specific inhibitor of MCD (CBM-301106), which we hypothesized would result in inhibition of CPT-I, reduction in fatty acid oxidation, a reciprocal activation of glucose oxidation, and diminished lactate production during demand-induced ischemia. Under normal-flow conditions, treatment with the MCD inhibitor significantly reduced oxidation of exogenous fatty acids by 82%, shifted the relationship between arterial fatty acids and fatty acid oxidation downward, and increased glucose oxidation by 50%. Ischemia was induced by a 20% flow reduction and β-adrenergic stimulation, which resulted in myocardial lactate production. During ischemia MCD inhibition elevated malonyl-CoA content fourfold, reduced free fatty acid oxidation rate by 87%, and resulted in a 50% decrease in lactate production. Moreover, fatty acid oxidation during ischemia was inversely related to the tissue malonyl-CoA content ( r = −0.63). There were no differences between groups in myocardial ATP content, the activity of pyruvate dehydrogenase, or myocardial contractile function during ischemia. Thus modulation of MCD activity is an effective means of regulating myocardial fatty acid oxidation under normal and ischemic conditions and reducing lactate production during demand-induced ischemia.

Evidence of a malonyl-CoA-insensitive carnitine palmitoyltransferase I activity in red skeletal muscle

2002 ◽  
Vol 282 (5) ◽  
pp. E1014-E1022 ◽  
Author(s):  
Jong-Yeon Kim ◽  
Timothy R. Koves ◽  
Geng-Sheng Yu ◽  
Tod Gulick ◽  
Ronald N. Cortright ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Splice Variants ◽  
Fiber Type ◽  
Carnitine Palmitoyltransferase ◽  
Acid Oxidation ◽  
Palmitate Oxidation ◽  
Oxidation Rates ◽  
Malonyl Coa ◽  
Carnitine Palmitoyltransferase I ◽  
Cpt I

Carnitine palmitoyltransferase I (CPT I), which is expressed as two distinct isoforms in liver (α) and muscle (β), catalyzes the rate-limiting step in the transport of fatty acid into the mitochondria. Malonyl-CoA, a potent inhibitor of CPT I, is considered a key regulator of fatty acid oxidation in both tissues. Still unanswered is how muscle β-oxidation proceeds despite malonyl-CoA concentrations that exceed the IC50 for CPT Iβ. We evaluated malonyl-CoA-suppressible [14C]palmitate oxidation and CPT I activity in homogenates of red (RG) and white (WG) gastrocnemius, soleus (SOL), and extensor digitorum longus (EDL) muscles. Adding 10 μM malonyl-CoA inhibited palmitate oxidation by 29, 39, 60, and 89% in RG, SOL, EDL, and WG, respectively. Thus malonyl-CoA resistance, which correlated strongly (0.678) with absolute oxidation rates (RG > SOL > EDL > WG), was greater in red than in white muscles. Similarly, malonyl-CoA-resistant palmitate oxidation and CPT I activity were greater in mitochondria from RG compared with WG. Ribonuclease protection assays were performed to evaluate whether our data might be explained by differential expression of CPT I splice variants. We detected the presence of two CPT Iβ splice variants that were more abundant in red compared with white muscle, but the relative expression of the two mRNA species was unrelated to malonyl-CoA resistance. These results provide evidence of a malonyl-CoA-insensitive CPT I activity in red muscle, suggesting fiber type-specific expression of distinct CPT I isoforms and/or posttranslational modulations that have yet to be elucidated.

Characterization of cardiac malonyl-CoA decarboxylase and its putative role in regulating fatty acid oxidation

1998 ◽  
Vol 275 (6) ◽  
pp. H2122-H2129 ◽  
Author(s):  
Jason R. B. Dyck ◽  
Amy J. Barr ◽  
Rick L. Barr ◽  
Pappachan E. Kolattukudy ◽  
Gary D. Lopaschuk
Keyword(s):  
Fatty Acid ◽  
Fatty Acid Oxidation ◽  
Fatty Acid Uptake ◽  
Partial Purification ◽  
Acid Oxidation ◽  
Acid Uptake ◽  
Adult Rats ◽  
Adult Rat ◽  
Oxidation Rates ◽  
Malonyl Coa

Malonyl-CoA is a potent inhibitor of fatty acid uptake into the mitochondria. Although the synthesis of malonyl-CoA in the heart by acetyl-CoA carboxylase (ACC) has been well characterized, no information is available as to how malonyl-CoA is degraded. We demonstrate that malonyl-CoA decarboxylase (MCD) activity is present in the heart. Partial purification revealed a protein of ∼50 kDa. The role of MCD in regulating fatty acid oxidation was also studied using isolated, perfused hearts from newborn rabbits and adult rats. Fatty acid oxidation in rabbit hearts increased dramatically between 1 day and 7 days after birth, which was accompanied by a decrease in both ACC activity and malonyl-CoA levels and a parallel increase in MCD activity. When adult rat hearts were aerobically reperfused after a 30-min period of no-flow ischemia, levels of malonyl-CoA decreased dramatically, which was accompanied by a decrease in ACC activity, a maintained MCD activity, and an increase in fatty acid oxidation rates. Taken together, our data suggest that the heart has an active MCD that has an important role in regulating fatty acid oxidation rates.

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