Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:6.4.1.2 (acetyl-CoA carboxylase)
2,876 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. The activity of acetyl-CoA carboxylase (EC 6.4.1.2) in extracts of freeze-clamped liver samples from fed or 24 h-starved virgin, pregnant, lactating and weaned rats was measured (i) immediately after preparation of extracts (;I activity'), (ii) after incubation of extracts with partially purified preparations of either rabbit muscle protein phosphatase 1 [Antoniw, Nimmo, Yeaman & Cohen (1977) Biochem. J.162, 423-433] or rabbit liver phosphatase [Brandt, Capulong & Lee (1975) J. Biol. Chem.250, 8038-8044] (;A activity') and (iii) after incubation with 20mm-potassium citrate before or after incubation with phosphatases (;C activity'). 2. Incubation of liver extracts at 30 degrees C without any additions resulted in activation of acetyl-CoA carboxylase that was shown to be due to dephosphorylation of the enzyme by endogenous protein phosphatase activity. This latter activity was not stimulated by Ca(2+) and/or Mg(2+) but was stimulated by 1 mm-Mn(2+). Incubation of extracts with either of the partially purified phosphatases (0.2-0.5 unit) resulted in faster dephosphorylation and activation. The activity achieved after incubation with either of the exogenously added phosphatases was similar. 3. The A and C activities increased during late pregnancy, were lower than in the virgin rat liver during early lactation and increased by 2-fold in liver of mid-lactating rats. Weaning of mid-lactating rats for 24 h resulted in no change in A and C activities but after 48 h weaning they were significantly lower than those in livers from suckled mothers. 4. The I activity followed a similar pattern of changes as the A and C activities during pregnancy and lactation such that, although the I/A and I/C activity ratios tended to be lower during late pregnancy and early lactation, there were no significant changes in I/A and I/C ratios between lactating and virgin animals. However, these ratios were significantly higher in liver from fed 24 h-weaned animals. 5. Starvation (24 h) resulted in a marked decrease in I activity for all animals studied except early-lactating rats. This was due to a combination of a decrease in the concentration of acetyl-CoA carboxylase in liver of starved animals (A and C activities) and a decrease in the fraction of the enzyme in the active form (lower I/C and I/A ratios). The relative importance of the two forms of regulation in mediating the starvation-induced fall in I activity was about equal in livers of virgin, pregnant and lactating animals. However, the decrease in I/A and I/C ratios was of dominating importance in livers of weaned animals. The A/C activity ratios were the same for livers from all animals studied. 6. The maximal activity of fatty acid synthase was also measured in livers and was highly and positively correlated with the A and C activities of acetyl-CoA carboxylase, suggesting that the concentrations of the two enzymes in the liver were controlled coordinately. 7. It is suggested that the lack of correlation between plasma insulin levels and rates of lipogenesis in the transition from the virgin to the lactating state may be explained by different effects of insulin and prolactin on the concentration of acetyl-CoA carboxylase in the liver and on the fraction of the enzyme in the active form.
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PMID:Changes in the proportion of acetyl-CoA carboxylase in the active form in rat liver. Effect of starvation, lactation and weaning. 612 71

Acetyl-CoA carboxylase phosphatase has been purified from the rat epididymal fat pad. The phosphatase occurs in a complex with the carboxylase. In the purification of the phosphatase, the high molecular weight complex was initially separated by sucrose gradient centrifugation, and the phosphatase was isolated from the complex by adjusting to 80% saturation with ethanol and by chromatography on Sephadex G-75. The molecular weight of the phosphatase is 71,000 as determined by sodium dodecyl sulfate gel electrophoresis and gel chromatography on Sephacryl-200 in the presence of 6 M urea. The Km for acetyl-CoA carboxylase and glycogen phosphorylase a are 1.5 microM and 37 microM, respectively. The phosphatase has a broad substrate specificity, being active toward glycogen synthase, 3-hydroxy-3-methylglutaryl-CoA reductase, phosphorylase a, phosphoprotamine, and p-nitrophenyl phosphate, in addition to acetyl-CoA carboxylase from fat tissue and liver. Acetyl-CoA carboxylase inhibits the dephosphorylation of phosphoprotamine, indicating that the same activity is responsible for dephosphorylating both substrates. The phosphatase requires no metal ion for activity and is not inhibited by the rat liver phosphorylase phosphatase inhibitor protein. The significance of these findings is discussed in relation to the regulation of acetyl-CoA carboxylase, and the phosphatase is compared to other phosphoprotein phosphatases.
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PMID:Purification and properties of acetyl-CoA carboxylase phosphatase. 625 18

The MgATP-dependent phosphorylase phosphatase was found to have a broad substrate specificity. Its activity against all phosphoproteins tested was dependent upon preincubation with the activating factor FA and MgATP. The enzyme dephosphorylated and inactivated phosphorylase kinase and inhibitor 1, and dephosphorylated and activated glycogen synthase and acetyl-CoA carboxylase. Glycogen synthase was dephosphorylated at similar rates whether it had been phosphorylated by cyclic-AMP-dependent protein kinase, phosphorylase kinase or glycogen synthase kinase 3. The enzyme also catalysed the dephosphorylation of ATP citrate lyase, initiation factor eIF-2, and troponin I. The properties of the MgATP-dependent protein phosphatase from either dog liver or rabbit skeletal muscle showed a remarkable similarity to highly purified preparations of protein phosphatase 1 from rabbit skeletal muscle. The relative activities of the two enzymes against all phosphoproteins tested was very similar. Both enzymes dephosphorylated the beta-subunit of phosphorylase kinase 40-fold faster than the alpha-subunit, and both enzymes were inhibited by identical concentrations of the two proteins termed inhibitor 1 and inhibitor 2, which inhibit protein phosphatase 1 specifically. These results demonstrate that the MgATP-dependent protein phosphatase is a type-1 protein phosphatase, and is distinct from type-2 protein phosphatases which dephosphorylate the alpha-subunit of phosphorylase kinase and are unaffected by inhibitor 1 and inhibitor 2. The possibility that the MgATP-dependent protein phosphatase is an inactive form of protein phosphatase 1 and that both proteins share the same catalytic subunit is discussed.
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PMID:The MgATP-dependent protein phosphatase and protein phosphatase 1 have identical substrate specificities. 626 81

Methods were developed for quantifying protein phosphatases-1, 2A, 2B and 2C in cell extracts, and these procedures were exploited to determine their tissue and subcellular distributions. In addition, the contribution of each enzyme to the total protein phosphatase activity in skeletal muscle and liver extracts towards nine proteins involved in the control of glycogen metabolism, glycolysis/gluconeogenesis, fatty acid synthesis and cholesterol synthesis was assessed. Each protein phosphatase was present at significant concentrations in skeletal muscle, heart muscle, liver, brain and adipose tissue, although the relative amounts differed considerably. In skeletal muscle, protein phosphatase-1 was the major enzyme acting on phosphorylase, glycogen synthase and phosphorylase kinase (beta-subunit), and thus was the major protein phosphatase responsible for the inactivation of glycogenolysis and stimulation of glycogen synthesis. This idea was reinforced by the observation that 50% of the protein phosphatase-1 activity was associated with the protein-glycogen complex. In the liver, protein phosphatases-1, 2A and 2C each appear to play a role in the regulation of glycogen metabolism. Protein phosphatase-1 accounted for a significant fraction of the total potential activity towards phosphorylase and glycogen synthase, and was the major phosphorylase kinase (beta-subunit) phosphatase of this tissue. In addition, it was the only protein phosphatase present in the protein-glycogen complex. Protein phosphatase 2A was also a major phosphorylase phosphatase and glycogen synthase phosphatase in this tissue. Protein phosphatase 2C was a significant glycogen synthase phosphatase in the liver, but had negligible activity toward phosphorylase or phosphorylase kinase (beta-subunit). In the absence of Ca2+, protein phosphatase 2A was the major phosphorylase kinase (alpha-subunit) phosphatase and the only inhibitor-1 phosphatase, in skeletal muscle or liver. In the presence of Ca2+, protein phosphatase 2B accounted for most of the activity towards these substrates. Protein phosphatase 2A was the major enzyme acting on L-pyruvate kinase, ATP-citrate lyase and acetyl-CoA carboxylase in rat liver, suggesting an important role in the regulation of glycolysis/gluconeogenesis and fatty acid synthesis. Protein phosphatase 2C was the major enzyme acting on hydroxymethylglutaryl-CoA (HMG-CoA) reductase and HMG-CoA reductase kinase, suggesting an important role in the regulation of cholesterol synthesis. However, the observation that 20% of the protein phosphatase-1 in liver was associated with the microsomal fraction suggests that this enzyme may also be involved in regulating HMG-CoA reductase, which is tightly associated with microsomes. The activity of protein phosphatase-1 in dilute skeletal muscle and liver extracts was just as sensitive to inhibitor-1 and inhibitor-2 as the purified enzyme. In concentrated extracts, higher concentrations of the inhibitor proteins were required and the inhibition was time-dependent...
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PMID:The protein phosphatases involved in cellular regulation. 6. Measurement of type-1 and type-2 protein phosphatases in extracts of mammalian tissues; an assessment of their physiological roles. 630 29

An acetyl-CoA carboxylase has been purified from rat hindlimb muscle using ammonium sulfate fractionation and avidin-Sepharose affinity chromatography. SDS/PAGE of the isolated enzyme showed a major protein band at approximately 272 kDa and a minor band at 265 kDa. The liver acetyl-CoA carboxylase gave a major protein band at 265 kDa and a minor band at 280 kDa. Adipose tissue acetyl-CoA carboxylase migrated to the 265-kDa position on the gel. Western blots performed using streptavidin-alkaline-phosphatase suggest that the bands from the three tissues contain biotin. The present study has characterized the muscle and adipose tissue enzymes under steady-state kinetics and determined Michaelis constants for the substrates. The activation constant for citrate, an essential activator for both preparations, was 2.13 +/- 0.05 mM for the muscle enzyme and 3.02 +/- 0.12 mM for adipose tissue (P < 0.01). The Km values for the muscle acetyl-CoA carboxylase compared to the adipose tissue acetyl-CoA carboxylase were: ATP, 57.6 +/- 0.9 microM compared to 106.5 +/- 2.6 microM, P < 0.01; acetyl-CoA, 31.7 +/- 1.5 microM compared to 21.5 +/- 1.0 microM, P < 0.01; bicarbonate, 2.25 +/- 0.10 mM compared to 2.73 +/- 0.29 mM, P > 0.05. The muscle acetyl-CoA carboxylase was inhibited by malonyl-CoA (Ki = 10.6 +/- 1.0 microM) and palmitoyl-CoA (Ki = 2.2 +/- 0.3 microM). These properties are consistent with the hypothesis that regulation of acetyl-CoA carboxylase plays an important role in governing the rate of fatty acid oxidation in the skeletal muscle.
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PMID:Purification and characterization of rat skeletal muscle acetyl-CoA carboxylase. 762 70

The activation of hepatic glycogen synthase by the amino-acid-induced cell swelling has been attributed to the stimulation of [glycogen-synthase]-phosphatase resulting from an increase in the intracellular content in glutamate and aspartate, and a decrease in intracellular Cl-, which is a compensatory response to cell swelling [Meijer, A. J., Baquet, A., Gustafson, L., van Woerkom, G. M. & Hue, L. (1992) J. Biol. Chem. 267, 5823-5828]. Here we studied whether the activation of acetyl-CoA carboxylase by cell swelling could be explained by the same mechanism. The activation of endogenous or purified acetyl-CoA carboxylase was measured in gel-filtered liver extracts or cytosols. No activation could be observed under basal conditions but a fivefold stimulation was obtained with concentrations of glutamate (20-25 mM) found in hepatocytes incubated with glutamine. A similar stimulation was also observed with other dicarboxylic acids such as malonate and succinate, or with metal ions like Mg2+, Ca2+ and Mn2+ (10 mM). The addition of 50-100 mM Cl- was found to inhibit the activation of acetyl-CoA carboxylase by some 20-30%. Mg2+ was also found to stimulate the activation of the endogenous glycogen synthase. The glutamate-stimulated and Mg(2+)-stimulated activation of glycogen synthase and acetyl-CoA carboxylase was unaffected by 10 microM inhibitor-2, a specific inhibitory protein of protein phosphatase-1, but could be nearly completely blocked by the phosphatase inhibitor microcystin-LR. Our data suggest that the amino-acid-induced activation of acetyl-CoA carboxylase and glycogen synthase in the liver occurs by a common ionic mechanism.
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PMID:Mechanism of activation of liver acetyl-CoA carboxylase by cell swelling. 790 Oct 14

Acetyl-CoA carboxylase is regulated allosterically by citrate and covalently by a phosphorylation/dephosphorylation mechanism. We have isolated and purified from rat livers a novel kinase that phosphorylates and inactivates the carboxylase. This kinase is bound to the carboxylase and can be eluted in salt-rich solution. The native kinase exists as high molecular weight aggregates of a subunit that has a molecular weight of 40,000. The phosphorylation sites of the carboxylase were determined after tryptic and cyanogen bromide digestions of 32P-labeled carboxylase and separation of the peptides by various chromatographic procedures. Amino acid analyses of the phosphopeptides showed that the Ser77 and Ser1200 residues were the sites of phosphorylation. Treating the phosphorylated carboxylase with the Mn(2+)-dependent acetyl-CoA carboxylase phosphatase 2 removed the phosphate and reactivated the carboxylase. These results suggest that both this kinase and the acetyl-CoA carboxylase phosphatase 2 act at the same site(s) in the acetyl-CoA carboxylase molecule. Citrate dramatically inhibits the kinase-mediated phosphorylation of the carboxylase, suggesting that the allosteric modification and activation by citrate render the phosphorylation sites inaccessible to the kinase and therefore maintain high carboxylase activity. This observation indicates that there is a close interplay between the citrate effect on and phosphorylation of the carboxylase in regulating its activity.
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PMID:Isolation and characterization of a novel acetyl-CoA carboxylase kinase from rat liver. 790 95

The activation of hepatic acetyl-CoA carboxylase by Na(+)-cotransported amino acids such as glutamine has been attributed mainly to the stimulation of its dephosphorylation by accumulating dicarboxylic acids, e.g. glutamate. We report here on a hepatic species of protein phosphatase-2A that activates acetyl-CoA carboxylase in the presence of physiological concentrations of glutamate or Mg2+ and, under these conditions, accounts for virtually all the hepatic acetyl-CoA carboxylase phosphatase activity. Glutamate also stimulated the dephosphorylation of a synthetic pentadecapeptide encompassing the Ser-79 phosphorylation site of rat acetyl-CoA carboxylase, but did not affect the dephosphorylation of other substrates such as phosphorylase. Conversely, protamine, which stimulated the dephosphorylation of phosphorylase, inhibited the activation of acetyl-CoA carboxylase. A comparison with various species of muscle protein phosphatase-2A showed that the stimulatory effects of glutamate and Mg2+ on the acetyl-CoA carboxylase phosphatase activity are largely mediated by the regulatory A subunit. Glutamate and Mg2+ emerge from our study as novel regulators of protein phosphatase-2A when acting on acetyl-CoA carboxylase.
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PMID:Activation of hepatic acetyl-CoA carboxylase by glutamate and Mg2+ is mediated by protein phosphatase-2A. 864 8

Despite the high expression of 5'AMP activated protein kinase (AMPK) in heart, the activity and function of this enzyme in heart muscle has not been characterized. We demonstrate that rat hearts have a high AMPK activity, comparable to that found in liver, which could be stimulated up to 3-fold by 5'AMP. Cardiac AMPK is also under phosphorylation control, since in vitro incubation of cardiac AMPK with protein phosphatase 2A completely abolished activity, while incubation with ATP/Mg(2+) resulted in over a 2-fold increase in activity. To investigate the function of AMPK in heart muscle, isolated working rat hearts were subjected to 30 min of global no-flow ischemia, followed by 60 min of aerobic reperfusion. AMPK activity was increased in heart at the end of reperfusion compared to aerobic controls (379 +/- 53 (n=5) vs. 139 +/- 19 (n=5) pmol x min(-1) x mg protein(-1), P<0.05, respectively). Treatment of AMPK in vitro with protein phosphatase 2A reversed this activation. Since AMPK can phosphorylate and inactivate acetyl-CoA carboxylase (ACC) in other tissues, and heart ACC has an important role in regulating fatty acid oxidation, we measured ACC activity in hearts reperfused post-ischemia. ACC activity was decreased at the end of reperfusion compared to aerobic controls (3.64 +/- 0.36 (n=9) vs. 10.93 +/- 0.60 (n=11) nmol x min(-1) x mg protein(-1), respectively, P<0.05). A significant negative correlation (r= -0.78) was observed between AMPK activity and ACC activity measured in aerobic and reperfused ischemic hearts. Low ACC activity could be reversed if ACC was extracted from hearts in the absence of phosphatase inhibitors, suggesting that phosphorylation of ACC decreased enzyme activity. This suggests that following ischemia AMPK is phosphorylated and activated (possibly by an AMPK kinase). AMPK then phosphorylates and inactivates ACC. The resultant decrease in malonyl-CoA levels could explain the acceleration of fatty acid oxidation that is observed during reperfusion of ischemic hearts.
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PMID:Characterization of 5'AMP-activated protein kinase activity in the heart and its role in inhibiting acetyl-CoA carboxylase during reperfusion following ischemia. 865 52

When mouse 30A5 preadipocytes are exposed to high glucose concentrations, acetyl-CoA carboxylase is induced through glucose activation of promoter II of the acetyl-CoA carboxylase gene. Glucose treatment of the cells increases Sp1 binding to two GC-rich glucose response elements in promoter II. We have investigated the mechanism by which glucose increases Sp1 binding and transactivation of promoter II in 30A5 cells. DNA mobility shift assays have shown that nuclear extracts from glucose-treated cells exhibit increased Sp1 binding activity. This increase in the binding activity is not due to glucose-mediated changes in the amount of Sp1 in the nucleus but to an increase in the activity that modifies Sp1 so that it binds more effectively to the promoter sequence. This Sp1 modifying activity is inhibited by okadaic acid and phosphatase inhibitor 2, and has a molecular mass of 38-42 kDa. The catalytic subunit of type 1 protein phosphatase, whose molecular mass is 38 kDa, also increased the ability of Sp1 to bind to promoter II. Treatment of nuclear extract with antibodies against the catalytic subunit partially suppressed the nuclear activity for Sp1 activation. From these results, we conclude that the Sp1 transcription factor exhibits enhanced binding to promoter II and transcriptional activation is the result of glucose-induced dephosphorylation by type 1 phosphatase.
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PMID:Dephosphorylation of Sp1 by protein phosphatase 1 is involved in the glucose-mediated activation of the acetyl-CoA carboxylase gene. 866 83


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