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 effects of the protein phosphatase inhibitors okadaic acid and microcystin LR on the regulation by insulin of pyruvate dehydrogenase and acetyl-CoA carboxylase have been studied in rat epididymal fat-pads and isolated cells. These inhibitors both completely blocked the phosphatase activity (against phosphorylase a) present in extracts of epididymal fat-pads, with half-maximal effects in the nanomolar range. 2. Okadaic acid treatment of pads and cells lowered the activity of acetyl-CoA carboxylase assayed in tissue extracts, both before and after treatment of the extracts with the activator, citrate. Further, okadaic acid treatment abolished the 2-3-fold difference in activity observed between extracts from control and insulin-treated tissues, assayed without prior treatment with citrate. 3. Incubation of pads with [32P]Pi, sufficient to label the intracellular pool of ATP, demonstrated that okadaic acid increased the overall phosphorylation of acetyl-CoA carboxylase on a number of distinct sites, as judged by two-dimensional mapping of tryptic peptides. These included the 'I-peptide' [Brownsey & Denton (1982) Biochem. J. 202, 77-86], the phosphorylation of which may be associated with the stimulation of the activity of the enzyme by insulin, as well as inhibitory phosphorylation sites. 4. Incubation with 1 microM-okadaic acid had no effect on the basal level of active pyruvate dehydrogenase apparent after tissue extraction, but abolished the 2-3-fold increase in this parameter which was elicited by insulin in the absence of okadaic acid. However, okadaic acid treatment did not affect the persistent increase in active pyruvate dehydrogenase levels which was apparent in mitochondria subsequently isolated from insulin-treated pads and re-incubated with an oxidizable substrate. It is concluded that the effects of okadaic acid are exerted through changes in metabolite concentrations rather than some direct action on the signalling pathway whereby insulin stimulates pyruvate dehydrogenase. 5. Microcystin LR did not mimic the effects of okadaic acid on intact cells and pads described above.
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PMID:Effects of protein phosphatase inhibitors on the regulation of insulin-sensitive enzymes within rat epididymal fat-pads and cells. 167 87

The role of cyclic AMP in acute regulation of the metabolism of mammary tissue in the lactating rat was examined by measuring the activity ratio of cyclic AMP-dependent protein kinase (A-kinase) and by examining the properties of this enzyme in its two major isoenzymic forms. Isoenzyme II is the major form in soluble extracts of rat mammary tissue. A-kinase activity ratio in such extracts is unaffected by starvation of the lactating rat. Treatment of the intact rat with isoprenaline, or addition of isoprenaline to incubations in vitro of mammary acini, resulted in a major increase in the activity ratio of A-kinase. These treatments equally affected isoenzymes I and II. The treatment in vitro lead to a rapid depletion of A-kinase as subsequently measured in extracts of acini. The degree of activation of the enzymes acetyl-CoA carboxylase and glycogen phosphorylase in extracts of mammary tissue and of acini was assessed as a function of these treatments. The increased activation of A-kinase induced by isoprenaline was unaccompanied by significant changes in the activity of acetyl-CoA carboxylase in acini, although we previously showed that this agent activates acetyl-CoA carboxylase in intact mammary tissue. Contrastingly, isoprenaline-induced enhancement of A-kinase activity was accompanied by an increase in the activity ratio of phosphorylase in acini. These results indicate that: (a) a normal response of expressed A-kinase activity to cyclic AMP operates in mammary acini and mammary tissue from lactating rats; (b) rapid modulation of the total amount of soluble A-kinase is mediated in mammary epithelial cells by cyclic AMP; (c) phosphorylase, an ultimate target of the protein phosphorylation cascade initiated by A-kinase, is activated in acini under conditions where A-kinase activity is enhanced; and (d) mechanisms other than that of the A-kinase phosphorylation/inhibition model for acetyl-CoA carboxylase regulation must operate in mammary tissue preparations and in vivo to account for the response of this enzyme to enhanced A-kinase activity.
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PMID:Cyclic AMP-dependent protein kinase in mammary tissue of the lactating rat. Activity ratio and responsiveness of the target enzymes acetyl-CoA carboxylase and glycogen phosphorylase to beta-adrenergic stimulation. 196 34

Acetyl-CoA carboxylase was isolated from rat liver by polyethylene glycol precipitation and avidin affinity chromatography. Sodium dodecyl sulfate electrophoresis of the enzyme gives one protein band (Mr 250,000). Phosphate analysis of the carboxylase showed the presence of 8.3 mol of phosphate/mol of subunit (Mr 250,000). The purified carboxylase has low activity in the absence of citrate (specific activity = 0.3 units/mg). However, addition of 10 mM citrate activates the carboxylase 10-fold, with half-maximal activation observed at 2 mM citrate, well above the physiological citrate level. Using this carboxylase as a substrate, we have isolated from rat liver a protein that activates the enzyme about 10-fold. This protein has been purified to near homogeneity (Mr 90,000). Incubation of this protein with 32P-labeled acetyl-CoA carboxylase results in a time-dependent activation of carboxylase with concomitant release of 32Pi, indicating that this protein is a phosphoprotein phosphatase. Both activation and dephosphorylation are dependent on Mn2+, but not citrate. This phosphatase does not hydrolyze p-nitrophenyl phosphate but does show high affinity for acetyl-CoA carboxylase (Km = 0.2 microM) as compared to its action on phosphorylase a (Km = 5.5 microM) and phosphohistone (Km = 20 microM). Activated acetyl-CoA carboxylase was isolated after dephosphorylation by the phosphatase. Such preparations contain about 5 mol of phosphate/mol of subunit and have specific activities of 2.6-3.0 units/mg in the absence of citrate. These activities are comparable to those of the phosphorylated carboxylase in the presence of 10 mM citrate. Thus, dephosphorylation by the Mn2+-dependent phosphatase renders the carboxylase citrate-independent, as compared to the phosphorylated form, which is citrate-dependent. To our knowledge this is the first report of a preparation of animal acetyl-CoA carboxylase that has substantial catalytic activity independent of citrate.
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PMID:Activation of acetyl-CoA carboxylase. Purification and properties of a Mn2+-dependent phosphatase. 286 Jan 6

Acetyl-CoA carboxylase from liver exhibits a linear inverse relationship between the ratio of enzymic activities at 0 and 2 mM citrate and the extent of phosphorylation by its kinase, and this citrate activity ratio method was used to examine the effect of nutritional conditions on the phosphorylation state of the enzyme. This method showed that the calculated phosphorylation state, being the extent of phosphorylation at sites accessible to carboxylase kinase, was highest in the livers of starved rats, lower in those fed normally, and lower still in starved rats which had been refed for 48 h on a fat-free diet. The actual values were 0.44, 0.26, and 0 mol of P/subunit, respectively, provided that liver samples were frozen rapidly to liquid nitrogen temperatures and extracted with stopping buffers at temperatures well below freezing. Normal homogenization with stopping buffers (containing inhibitors for protein kinases and phosphatases) resulted in much higher calculated phosphorylation states. The effect of nutritional conditions on the phosphorylation state as estimated reported above was confirmed by purifying the carboxylase from livers of rats, measuring the amount of phosphate which could be incorporated by carboxylase kinase, and comparing this with the phosphorylation state calculated from the citrate activity ratio method or the specific activity. Furthermore, treatment with protein phosphatase of carboxylase from starved rats resulted in the largest increase in specific activity, that from the starved/refed rats in the least. Finally, the effects of hyperglycemia on carboxylase and phosphorylase characteristics in the livers of intact rats were ascertained by taking liver samples and preparing crude extracts by the rapid freezing method described above. Hyperglycemia caused a rapid increase in the activity of the carboxylase and a rapid decrease in its putative phosphorylation state as measured by the citrate activity ratio method. Phosphorylase was also dephosphorylated, as indicated by a decrease in phosphorylase a activity. We conclude that the citrate activity ratio method is a valid test for the phosphorylation state of acetyl-CoA carboxylase in crude extracts of tissue.
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PMID:Phosphorylation state of acetyl-coenzyme A carboxylase. II. Variation with nutritional condition. 287 34

Although glucocorticoid and thyroid hormones are known to act synergistically to stimulate surfactant production, they have opposite effects on other parameters of fetal lung maturation. We recently reported that the developmental increases in de novo fatty acid synthesis and glycogen accumulation in fetal rat lung were accelerated by dexamethasone but prevented by triiodothyronine and that the dexamethasone-induced increases were diminished when the two hormones were administered together. We have now examined the effects of maternal administration of these hormones on activities of enzymes of lung fatty acid synthesis and glycogen metabolism in the rat. There was a developmental increase in fatty-acid synthase activity between 19 and 21 days gestation. This activity was increased by dexamethasone but decreased by triiodothyronine. When the two hormones were administered together the stimulatory effect of dexamethasone was decreased from 56% to 29%. The stimulatory effect on fatty-acid synthase was also observed in fetal lung explants cultured in the presence of dexamethasone. This shows that the effect of the hormone was directly on the fetal lung. Dexamethasone had no effect on liver fatty-acid synthase. There was a developmental decrease in acetyl-CoA carboxylase activity but it was not affected by the hormones. These data show that the developmental and hormone-induced changes in fetal lung de novo fatty acid synthesis are mediated by fatty-acid synthase. Although there were developmental changes in fetal lung 6-phosphofructokinase, glycogen synthase and glycogen phosphorylase activities, these enzymes were not affected by the hormones.
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PMID:Effects of glucocorticoid and thyroid hormones on regulatory enzymes of fatty acid synthesis and glycogen metabolism in developing fetal rat lung. 288 79

The phosphorylase phosphatases in rat and rabbit liver cytosol that are markedly stimulated by histone H1, protamine and polylysine were identified as protein phosphatases-2A0, 2A1 and 2A2 by anion-exchange chromatography, gel-filtration and immunotitration experiments. Histone H1 and protamine also stimulated the dephosphorylation of phosphorylase kinase, glycogen synthase, fructose-1,6-bisphosphatase, pyruvate kinase, acetyl-CoA carboxylase and phenylalanine hydroxylase by phosphatases-2A1 and 2A2, and with several of these substrates activation was even more striking (20-100-fold) than that observed with phosphorylase (approximately 5-fold). Activation by basic polypeptides did not involve dissociation of these phosphatases to the free catalytic subunit. The dephosphorylation of phosphorylase by protein phosphatase-1 was suppressed by basic polypeptides, protamine and polylysine being the most potent inhibitors. However, the dephosphorylation of glycogen synthase, pyruvate kinase and acetyl-CoA carboxylase were markedly stimulated by histone H1 and protamine (2-13-fold). Consequently, with the appropriate substrates, protein phosphatase-1 can also be regarded as a basic-polypeptide-activated protein phosphatase. Heparin stimulated (1.5-2-fold) the dephosphorylation of phosphorylase by phosphatases-2A0 and 2A1, provided that Mn2+ was present, but phosphatase-2A2 and the free catalytic subunit of phosphatase-2A were unaffected. Heparin, in conjunction with Mn2+, also stimulated (1.5-fold) the dephosphorylation of glycogen synthase (labelled in sites 3 abc), phosphorylase kinase and phenylalanine hydroxylase by phosphatase-2A1, but not by phosphatase-2A2. By contrast, the dephosphorylation of phosphorylase and phosphorylase kinase by protein phosphatase-1 was inhibited by heparin. However, dephosphorylation of glycogen synthase and pyruvate kinase by phosphatase-1 was stimulated by this mucopolysaccharide. The studies demonstrate that basic proteins can be used to distinguish protein phosphatase-1 from protein phosphatase-2A, but only if phosphorylase is employed as substrate. Optimal differentiation of the two phosphatases is observed at 30 micrograms/ml protamine or at heparin concentrations greater than 150 microM.
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PMID:The protein phosphatases involved in cellular regulation. 1. Modulation of protein phosphatases-1 and 2A by histone H1, protamine, polylysine and heparin. 298 84

A rat liver cAMP-independent protein kinase that phosphorylates peptide b of ATP-citrate lyase (Ramakrishna, S., Pucci, D. L., and Benjamin, W. B. (1983) J. Biol. Chem. 258, 4950-4956) has been purified to apparent homogeneity. The molecular weight, determined by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, sucrose density gradient, and by gel filtration, was found to be 36,000. This protein kinase phosphorylates in vitro ATP-citrate lyase, acetyl-CoA carboxylase, and glycogen synthase and does not phosphorylate phosphorylase, phosphorylase kinase, histone, phosvitin, and casein. It has Fa (activity factor) activity stimulating the ATP X Mg-dependent phosphatase and is therefore named a multifunctional protein kinase. This kinase differs from glycogen synthase kinase-3 with regard to substrate specificity, kinetic parameters, and physicochemical properties.
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PMID:Cyclic nucleotide-independent protein kinase from rat liver. Purification and characterization of a multifunctional protein kinase. 404 96

Acetyl-CoA carboxylase from rat epididymal fat tissue is activated by incubation at 30 C in the absence of citrate or metal ions. This activation is accompanied by a corresponding loss of 32P from the labeled enzyme, and it is not blocked by the heat-stable phosphorylase phosphatase inhibitor proteins from rabbit muscle. We have succeeded in separating an activity which activates and dephosphorylates acetyl-CoA carboxylase from the carboxylase using polyethylene glycol-6000. These results suggest that the temperature-dependent activation of acetyl-CoA carboxylase in crude or partially purified preparations results from dephosphorylation of the carboxylase by bound phosphatase.
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PMID:Heat activation of rat epididymal fat tissue acetyl-coa carboxylase is due to dephosphorylation by its endogenous phosphatase. 611 34

A protein kinase which phosphorylates and inactivates acetyl-CoA carboxylase has been purified to apparent homogeneity from rat liver. The kinase was found to exist in two forms: bound to carboxylase in a complex or in a free form that is in different stages of aggregation over a wide range of molecular weights. The purification of the kinase involved first partial purification of acetyl-CoA carboxylase through polyethylene glycol precipitation and DEAE-cellulose chromatography. The kinase was then separated from acetyl-CoA carboxylase by Sepharose 2B chromatography. The molecular weight of the kinase subunit was 170,000 as determined by sodium dodecyl sulfate-gel electrophoresis. The incorporation of 1 mol of phosphate/mole of carboxylase subunit caused complete inactivation of the carboxylase. Acetyl-CoA carboxylase, inactivated by the kinase, can be dephosphorylated and reactivated when incubated with phosphorylase phosphatase. The Km values of the kinase for acetyl-CoA carboxylase and ATP are 90 nM and 20 microM, respectively. The kinase was found to be cyclic AMP-independent, but activated by CoA. The protein kinase can phosphorylate acetyl-CoA carboxylase, protamine, and histones, but could not act on hydroxymethylglutaryl-CoA reductase or phosphorylase b.
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PMID:Purification and properties of a kinase which phosphorylates and inactivates acetyl-CoA carboxylase. 612 Jan 70

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


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