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. Acetyl-CoA carboxylase was purified to homogeneity, in the presence of protein phosphatase inhibitors, from rat liver sampled without freeze-clamping. The enzyme was in a highly phosphorylated state (4.8 mol/subunit) of low specific activity, and could be dramatically reactivated by treatment with protein phosphatase-2A. Amino acid sequencing and fast-atom-bombardment mass spectrometry showed that the enzyme was phosphorylated in Ser79, Ser1200 and Ser1215, the three sites known to be phosphorylated in cell-free assays by the AMP-activated protein kinase. 2. The inactive enzyme could also be completely reactivated using a limited treatment with trypsin, which removes the N-terminal segment containing Ser79 and reduces the phosphate content to 3.5 mol/subunit. These results strengthen previous findings that it is phosphorylation at Ser79 by the AMP-activated protein kinase that is responsible for the inactivation, and not the phosphorylation of the 220-kDa core fragment (which contains Ser1200 and Ser1215). 3. Analysis of the phosphorylation state of Ser79 in acetyl-CoA carboxylase from rat liver showed that phosphorylation occurs post mortem if freeze-clamping is not used. The higher phosphorylation observed in extracts made without freeze-clamping correlates with a large increase in AMP and decrease in ATP (presumably caused by hypoxia during removal of the liver), and with increased activity of the AMP-activated protein kinase. These results provide a rational explanation for the post mortem phosphorylation events, and re-emphasize the point that rapid cooling of cells and tissues is essential when measuring the expressed activity of acetyl-CoA carboxylase (as well as 3-hydroxy-3-methylglutaryl-CoA reductase). 4. Using the freeze-clamping procedure, the ratio of 'expressed' activity (measured in the presence of protein phosphatase inhibitors) to 'total' activity (measured after complete dephosphorylation) of rat liver acetyl-CoA carboxylase showed a marked diurnal rhythm, changing from 50% in the active form in the middle of the dark period to less than 10% active in the middle of the light period. The very low activity in the light period was associated with a high level of phosphorylation in Ser79. This diurnal rhythm is very similar to that previously described for the phosphorylation of 3-hydroxy-3-methylglutaryl-CoA reductase, another substrate for the AMP-activated protein kinase. Neither the activity of the AMP-activated protein kinase nor the content of AMP, ADP or ATP changed between the dark or light periods.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Diurnal rhythm of phosphorylation of rat liver acetyl-CoA carboxylase by the AMP-activated protein kinase, demonstrated using freeze-clamping. Effects of high fat diets. 134 20

The effect of cholesterol diet on the rate of mevalonic acid biosynthesis from 1-14C acetyl-CoA, 2-14C malonyl-CoA and the incorporation of these substrates into sterols and bile acids in rabbit liver were studied. Simultaneously, the activities of 3-hydroxy-3-methylglutaryl-CoA reductase (HMG-CoA reductase) and acetyl-CoA carboxylase and the biosynthesis of fatty acids from acetyl-CoA and malonyl-CoA were measured. Hypercholesterolemia was found to be concomitant with the inhibition of acetyl-CoA carboxylase activity only in cell-free (700 g) and mitochondrial fractions and slightly decreased the incorporation of acetyl-CoA and malonyl-CoA into fatty acids in the postmitochondrial fraction. The HMG-CoA reductase activity in all subcellular fractions except for the postmicrosomal one was inhibited under these conditions. A significant decrease of acetyl-CoA incorporation and an increase in malonyl-CoA incorporation into mevalonic acid in all liver fractions except for microsomal one were observed in rabbits with hypercholesterolemia. These data provide evidence for the existence of two pathways of mevalonic acid synthesis from the above-said substrates that are differently sensitive to cholesterol. Cholesterol feeding resulted in a decreased synthesis of the total unsaponified fraction including cholesterol from acetyl-CoA, malonyl-CoA and mevalonic acid. The rate of incorporation of these substrates into lanosterol was unchanged. All the indicated substrates (acetyl-CoA, malonyl-CoA, mevalonic acid) are precursors of bile acid synthesis in rabbit liver. Cholesterol feeding and the subsequent development of hypercholesterolemia resulted in bile acid synthesis stimulation, preferentially in the formation of the cholic + deoxycholic acids from these precursors.
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PMID:[Formation of mevalonic acid, sterols and bile acids from [1-14C]acetyl-CoA and [2-14C]malonyl-CoA in the liver of rabbits with experimental hypercholesterolemia]. 288 84

The relationship between cell density and de novo synthesis of sterols and fatty acids has been studied in monolayer cultures of L-M cells grown in serum-free medium. Incorporation of radioactivity from [14C] acetate or 3H2O into sterols and fatty acids declined sharply as cultures approached stationary phase. The activities of 3-hydroxy-3-methylglutaryl-CoA reductase and 3-hydroxy-3-methylglutaryl-CoA synthase declined in conjunction with the decrease in sterol synthesis; however, the activity of acetoacetyl-CoA thiolase did not decrease until after sterol synthesis had begun to decline. The magnitude of the initial decline in reductase activity was not diminished when activation of latent enzyme activity was prevented by addition of fluoride to cell homogenates. The diminution in the rate of fatty acid synthesis at high cell density was accompanied by a decrease in the activity of fatty acid synthetase, whereas the activity of acetyl-CoA carboxylase increased slightly. The data suggest that lipogenesis is regulated in coordination with the changes in the rate of cell proliferation that occur when L-M cells attain a high density in monolayer culture. Moreover, these studies establish the feasibility of using the L-M cell culture system to investigate the relationship between cell density and the enzymatic regulation of lipogenesis.
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PMID:Changes in synthesis of sterols and fatty acids associated with inhibition of growth of L-M cells at high cell density. 611 18

The effects of Triton WR 1339, starvation and cholesterol diet on the activities of 3-hydroxy-3-methylglutaryl-CoA reductase (HMG-CoA reductase) and acetyl-CoA carboxylase and on the rates of mevalonic acid (MVA) biosynthesis from acetyl-CoA and malonyl-CoA in the soluble (140 000 g) and microsomal fractions of rat liver, on the rate of incorporation of these substrates into squalene, cholesterol and lanosterol in the rat liver postmitochondrial fraction and on the rate of fatty acid biosynthesis was studied. The administration of Triton WR 1339 (200 mg per 100 g of body weight twice) stimulated the activity of HMG-CoA reductase and MVA biosynthesis from acetyl-CoA and malonyl-CoA in the intact and solubilized microsomal fractions and had no effect on these parameters in the soluble fraction. Starvation for 36 hrs did not cause inhibition of the reductase activity or MVA biosynthesis from both substrates in the soluble fraction. Alimentary cholesterol significantly increased the activity of HMG-CoA reductase, had no effect on the rate of MVA biosynthesis from acetyl-CoA and stimulated the malonyl-CoA incorporation in to MVA in the soluble fraction. Starvation an alimentary cholesterol inhibited the HMG-CoA reductase activity and MVA biosynthesis from both substrates in the solubilized microsomal fraction. Triton WR 1339 stimulated 4--19-fold the lipid formation in the total unsaponified fraction and its components i.e. squalene, lanosterol, cholesterol, from acetyl-CoA and only insignificantly (1,2--1,7-fold) increased malonyl-CoA incorporation into these compounds. Starvation and alimentary cholesterol repressed lanosterol and cholesterol biosynthesis from acetyl-CoA, decreased malonyl-CoA incorporation into these sterols and had no influence on squalene biosynthesis from the two substrates. Triton WR 1339 and starvation inhibited the acetyl-CoA carboxylase activity, unaffected by alimentary cholesterol. No significant changes in the rate of fatty acid biosynthesis from the substrates were observed. The data obtained provide evidence for the existence of autonomic pathways of MVA biosynthesis localized in the soluble and microsomal fractions of rat liver. The pathway of MVA biosynthesis in the soluble fraction is less sensitive to regulatory factors. Sterol biosynthesis from malonyl-CoA is also more resistant to regulatory effects than sterol biosynthesis from acetyl-CoA. This suggests that HMG-CoA reductase localized in the soluble fraction takes part in MVA and sterol biosynthesis from malonyl-CoA.
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PMID:[Activities of 3-hydroxyl-3-methylglutaryl-CoA reductase and acetyl-CoA carboxylase and the rate of mevalonic acid, squalene, sterol and fatty acid biosynthesis from [1-14C]acetyl-CoA and [2-14C]malonyl-CoA in rat liver: effects of Triton WR 1339, starvation and cholesterol diet]. 611 54

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

The nature of the protein phosphatases involved in the regulation of glycolysis/gluconeogenesis, fatty acid synthesis and cholesterol synthesis in rat liver has been investigated using L-pyruvate kinase, ATP-citrate lyase, acetyl-CoA carboxylase and hydroxymethylglutaryl-CoA reductase as substrates. The results show that protein phosphatases-1, 2A and 2C are the only significant protein phosphatases in rat liver acting on these four substrates. The relationship of these three enzymes to other protein phosphatases described in the literature is discussed.
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PMID:The protein phosphatases involved in cellular regulation. 3. Fatty acid synthesis, cholesterol synthesis and glycolysis/gluconeogenesis. 630 26

The possibility of biosynthesis of cholic (I) and chenodeoxycholic (II) acids from [1-14C]acetyl-CoA and [2-14C]malonyl-CoA in a reconstituted system of rat liver and the incorporation of acetyl-CoA into these bile acids under conditions of acetyl-CoA carboxylase activation by citrate or its inhibition by avidin were studied. The effects of Triton WR 1339 and cholesterol feeding on acetyl-CoA and malonyl-CoA incorporation into I and II were investigated. Teh incorporation of both substrates into the total unsaponifiable lipid fraction and fatty acids was demonstrated. The reconstituted system of rat liver was found able to synthesize and I and II not only from acetyl-CoA, but from malonyl-CoA as well. The rate of malonyl-CoA incorporation into the bile acids was somewhat higher than that of acetyl-CoA incorporation. Preincubation of the reconstituted system with citrate stimulated the rate of acetyl-CoA incorporation into I. Stimulation of biosynthesis of I occurred independently of the diurnal rhythm of the 3-hydroxy-3-methylglutaryl-CoA reductase (HMG-CoA reductase) activity. An addition of avidin to the reconstituted system preincubated with citrate caused inhibition of acetyl-CoA incorporation both into fatty acids and into I. The rate of biosynthesis of II remained practically unchanged in both cases. Treatment with Triton WR 1339 had only a slight effect, while cholesterol feeding significantly stimulated the incorporation of acetyl-CoA and malonyl-CoA into I and II. The results obtained suggest the participation of malonyl-CoA in formation of bile acids, preferentially cholic acid, and in a lesser degree, in sterol biosynthesis. Data from stimulation of bile acid biosynthesis under cholesterol feeding suggest that HMG-CoA reductase localized in the soluble fraction of rat liver is involved in bile acid biosynthesis.
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PMID:[Biosynthesis of cholic and chenodeoxycholic acids from [1-14C]acetyl-CoA and [2-14C]malonyl-CoA in a reconstituted system from the rat liver]. 723 4

AMP-activated protein kinase is a multisubstrate protein kinase that, in liver, inactivates both acetyl-CoA carboxylase, the rate-limiting enzyme of fatty acid synthesis, and 3-hydroxy-3-methyl-glutaryl-CoA reductase, the rate-limiting enzyme of cholesterol synthesis. AICAR (5-amino 4-imidazolecarboxamide ribotide, ZMP) was found to stimulate up to 10-fold rat liver AMP-activated protein kinase, with a half-maximal effect at approximately 5 mM. In accordance with previous observations, addition to suspensions of isolated rat hepatocytes of 50-500 microM AICAriboside, the nucleoside corresponding to ZMP, resulted in the accumulation of millimolar concentrations of the latter. This was accompanied by a dose-dependent inactivation of both acetyl-CoA carboxylase and 3-hydroxy-3-methylglutaryl-CoA reductase. Addition of 50-500 microM AICAriboside to hepatocyte suspensions incubated in the presence of various substrates, including glucose and lactate/pyruvate, caused a parallel inhibition of both fatty acid and cholesterol synthesis. With lactate/pyruvate (10/1 mM), half-maximal inhibition was obtained at approximately 100 microM, and near-complete inhibition at 500 microM AICAriboside. These findings open new perspectives for the simultaneous control of triglyceride and cholesterol synthesis by pharmacological stimulators of AMP-activated protein kinase.
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PMID:Inhibition of fatty acid and cholesterol synthesis by stimulation of AMP-activated protein kinase. 773 63

The AMP-activated protein kinase is responsible for the regulation of fatty acid synthesis by phosphorylation of acetyl-CoA carboxylase. It may also regulate cholesterol synthesis via phosphorylation and inactivation of hormone-sensitive lipase and hydroxymethylglutaryl-CoA reductase. We have purified the AMP-activated protein kinase 14,000-fold from porcine liver. The 63-kDa catalytic subunit co-purifies with two proteins of 40 and 38 kDa that may function as subunits. Partial amino acid sequence of the 63-kDa subunit revealed a striking homology with the catalytic domain of the yeast protein kinase transcriptional regulator Snf1 and its plant homologs. The Snf1 (72 kDa) and Snf4 (36 kDa) complex was also purified and found to phosphorylate the AMP-activated protein kinase peptide substrate, HMRSAMSGLHLVKRR-amide, but was not activated by AMP. Both Snf1/4 and the AMP-activated protein kinase phosphorylate and inactivate yeast acetyl-CoA carboxylase in vitro. These results indicate that during evolution the catalytic domain sequences of the Snf1 protein kinase subfamily have been exploited in the control of mammalian lipid metabolism and raise the possibilities that the AMP-activated protein kinase may have other substrates involved in regulating gene expression pathways, as well as Snf1 homologs participating in the control of lipid metabolism in many eukaryotic organisms.
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PMID:Mammalian AMP-activated protein kinase shares structural and functional homology with the catalytic domain of yeast Snf1 protein kinase. 790 77


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