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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)

The isolation and biochemical properties of a Saccharomyces cerevisiae mutant (acc1-167) defective in acetyl-CoA carboxylase [acetyl-CoA:carbon-dioxide ligase (ADP forming), EC 6.4.1.2] activity are described. The mutant is deficient in de novo biosynthesis of long-chain fatty acids and specifically requires a saturated fatty acid of chain length 14-16 C atoms for growth. Fatty acid synthetase levels were normal, but the acetyl-CoA carboxylase specific activity of the purified enzyme was reduced to approximately 5% compared to wild-type yeast. Upon sodium dodecyl sulfate/polyacrylamide gel electrophoresis the purified mutant enzyme migrated as a single band and was essentially indistinguishable from the wild-type enzyme. The study of acetyl-CoA carboxylase partial activities revealed that the biotin incorporation capacity and the transcarboxylase partial activity were unaffected whereas the biotin carboxylase component enzyme exhibited less than 10% of wild-type specific activity. This biotin carboxylase mutational deficiency could be ascribed to a more than 90% reduction of Vmax and to a comparable increase in the Km value for ATP, which was accompanied by an increased requirement for Mg2+. It is concluded that acc1-167 contains a structural gene mutation in the biotin carboxylase domain of acetyl-CoA carboxylase.
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PMID:Fatty acid-requiring mutant of Saccharomyces cerevisiae defective in acetyl-CoA carboxylase. 610 40

Acetyl-CoA carboxylase [acetyl-CoA:carbon-dioxide ligase (ADP-forming), EC 6.4.1.2] is activated by physiological concentrations of CoA. The CoA concentration dependency of this activation is sigmoidal; below 60 microM there is little or no activation, but the activation observed between 60 and 120 microM indicates that small changes in the concentration of CoA can cause significant changes in carboxylase activity. CoA activation of acetyl-CoA crboxylase accompanies polymerization of acetyl-CoA carboxylase. However, the binding site for CoA appears to be different from that of citrate. In contrast to citrate activation, which changes only the Vmax of the reaction, CoA activation of carboxylase results in polymeric forms with a lower Km for acetyl-CoA. The Km for acetyl-CoA is 0.4 mM in the control enzyme, whereas that of the CoA-activated enzyme is as low as 4 microM. The Km for ATP was not changed. Derivatives of CoA were not effective in activating the carboxylase, indicating that the CoA effect is specific. Arguments are presented that CoA could be a physiologically significant positive effector of the carboxylase.
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PMID:Regulation of acetyl-coA carboxylase: properties of coA activation of acetyl-coA carboxylase. 610 89

Acetyl-CoA carboxylase has been purified from lactating rat mammary gland using a combination of ammonium sulphate and poly(ethyleneglycol) precipitations. The enzyme was purified from 35--70-fold with a yield of over 50%, the exact figures being difficult to estimate because of activation of the enzyme that occurs during the preparation. The preparation was homogeneous by the criterion of polyacrylamide gel electrophoresis in sodium dodecyl sulphate and had a single subunit of molecular weight 240,000, containing 1.02 +/- 0.04 molecules of biotin and 3.1 +/- 1.7 molecules of alkali-labile phosphate per subunit. The purified enzyme was phosphorylated and inactivated rapidly when incubated in the presence of [gamma 32P]ATP and magnesium ions with the purified catalytic subunit of cyclic-AMP-dependent protein kinase from rabbit skeletal muscle. Both phosphorylation and inactivation are blocked by the heat-stable protein inhibitor of cyclic-AMP-dependent protein kinase, and can be reversed by incubation with purified protein phosphatase-1 from rabbit skeletal muscle. The inactivation by the protein kinase and reactivation by the protein phosphatase correlate with the near-stoichiometric phosphorylation and dephosphorylation of site(s) located in a single tryptic peptide. Phosphorylation does not affect the Km for substrates, but brings about a twofold decrease in V and a twofold increase in the apparent dissociation constant for the allosteric activator, citrate. We also present evidence that the activation of rabbit mammary acetyl-CoA carboxylase by protein phosphatase-1 described previously [Hardie and Cohen (1979) FEBS Lett. 103, 333-338] is due to dephosphorylation at site(s) which are not phosphorylated by either cyclic-AMP-dependent protein kinase or acetyl-CoA carboxylase kinase-2. These results suggest that the rapid inactivation of acetyl-CoA carboxylase, and hence fatty acid synthesis, by adrenaline in adipose tissue, or glucagon in the liver, is due to phosphorylation of the enzyme by cyclic-AMP-dependent protein kinase.
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PMID:Reversible phosphorylation and inactivation of acetyl-CoA carboxylase from lactating rat mammary gland by cyclic AMP-dependent protein kinase. 610 9

Among more than 7000 mutants of Saccharomyces cerevisiae, requiring saturated fatty acids, 61 acetyl-CoA-carboxylase-deficient strains have been identified. According to their mutual complementation characteristics these mutants have been assigned to two different genes, acc1 and acc2. Both acetyl-CoA carboxylase genes are unlinked to each other and to the fatty acids synthetase genes fas1 and fas2. The acetyl-CoA carboxylases of several acc1 and acc2 mutants have been purified and assayed for their overall and component enzyme activities. Besides overall acetyl-CoA carboxylation, which was lost in all cases, both component enzymes, biotin carboxylase and transcarboxylase, were simultaneously affected in most mutants, though often to a different relative extent. Similarly, the comparison of biochemical and genetic complementation data revealed no basis for a clear distinction between specific biotin carboxylase and transcarboxylase mutants. These results suggest that acc1 is a cluster gene coding for a multifunctional protein harboring both acetyl-CoA carboxylase component enzyme activities on the same polypeptide chain. The acetyl-CoA carboxylase isolated from acc2 mutants was free of biotin. Correspondingly, biotin:apoacetyl-CoA-carboxylase ligase activity was missing in acc2 mutants. Therefore, it is concluced that the primary defect in acc2 mutants is in the biotin:apocarboxylase ligase. In agreement with this conclusion, the acc2 acetyl-CoA carboxylase can be activated, in the presence of biotin and ATP, by ligase preparations from wild-type or acc1 mutant cells. By the use of these mutants, evidence was obtained that in vivo the biotinylation of both acetyl-CoA carboxylase and pyruvate carboxylase is catalyzed by the same ligase.
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PMID:Yeast mutants defective in acetyl-coenzyme A carboxylase and biotin: apocarboxylase ligase. 610 18

Age-related changes in the activity of rat liver acetyl-CoA carboxylase and the phospholipid content have been investigated in detail between birth and the age of 2.5 years. During postnatal development and ageing the enzyme shows varying kinetic properties as well as different specific activities. During the first days of postnatal life the activity declines. A minimum of 20% from adult levels is found 4 days after birth. Thereafter the specific activity rapidly increases. The highest specific activities are found in old animals. Major changes can be found in the kinetics of the activation of the enzyme by citrate. The influence of ATP concentration shows minor age-related alterations. The phospholipid composition of the rat liver is age-dependent, too. The coincidence of modifications in the phospholipid composition and correlated variations of the acetyl-CoA carboxylase activity as well as developmental increase of the endoplasmic reticulum in the liver and the growth rate of the rat brain are discussed.
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PMID:Age-related changes in the kinetic properties of the acetyl-CoA carboxylase and phospholipid content of rat liver. 611 Aug 11

Fatty acid synthesis is traditionally viewed as being confined to the cytosolic cellular fraction, although a substantial body of data indicates that both microsomes and mitochondria are capable of initiating fatty acid synthesis and may contain acetyl-CoA carboxylase [acetyl-CoA:carbon-doxide ligase (ADP-forming), EC 6.4.1.2], fatty acid synthetase, and ATP-citrate lyase [ATP citrate (pro-3S)-lyase; ATP:citrate oxaloacetate-lyase (pro-3S-CH2COO- leads to acetyl-CoA; ATP-dephosphorylating), EC 4.1.3.8] activities. We have identified 32P-labeled acetyl-CoA carboxylase and 32P-labeled ATP-citrate lyase by immunoprecipitation of a rat hepatocyte microsomal preparation. In the transition between the fasting state (low rates of lipogenesis) and fasting/re-feeding (high rates), the fraction of total cytosolic plus microsomal acetyl-CoA carboxylase in the microsomes increases from 6% to 43%, whereas the microsomal proportion of total fatty acid synthetase and ATP-citrate lyase remains approximately 10%. Microsome isolation conditions favoring carboxylase polymerization (presence of citrate) promote microsomal association, whereas conditions favoring enzyme protomerization (malonyl-CoA, preincubation with cyclic AMP/ATP/Mg2+) diminish this association. The microsomal enzyme has a 5-fold higher specific activity than the cytosolic enzyme as determined by immunotitration. Sucrose density gradient analysis of the microsomal fraction indicates that a substantial portion of carboxylase activity sediments with marker enzymes for endoplasmic reticulum, plasma membrane, Golgi apparatus, and outer mitochondrial membrane, while cytosolic enzyme or isolated enzyme incubated under polymerizing conditions does not penetrate the gradient. These data suggest that the microsomes may be a significant locus of fatty acid synthesis initiated with association of acetyl-CoA carboxylase polymer with this fraction.
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PMID:Microsomal acetyl-CoA carboxylase: evidence for association of enzyme polymer with liver microsomes. 611 83

Avidin affinity chromatography was used to rapidly purify acetyl-CoA carboxylase to homogeneity in high yield from chicken liver. Dissociation of the purified carboxylase with dodecyl sulfate yielded a single size class of subunit polypeptide of 225,000 daltons. A steady state kinetic analysis of the carboxylase-catalyzed carboxylation of acetyl-CoA gave rise to intersecting line patterns in all double-reciprocal plots of initial velocity with each substrate pair, i.e. ATP . Mg and HCO3(-) and acetyl-CoA. It was concluded that the kinetic mechanism involves a quaternary complex of the enzyme, ADP, Pi, and acetyl-CoA rather than a double displacement as previously believed. The ordered addition of ATP, HCO3(-), and then acetyl-CoA, to the citrate-activated form of the carboxylase is the kinetic mechanism most consistent with the results.
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PMID:Acetyl coenzyme A carboxylase. Rapid purification of the chick liver enzyme and steady state kinetic analysis of the carboxylase-catalyzed reaction. 611 14

The activities of acetyl-CoA carboxylase, ATP citrate-lyase and fatty acid synthetase remained low until parturition at 22 days of gestation and increased significantly within 1 day post partum. Administration of progesterone on days 20 and 21 and at parturition abolished the increases for at least 48 h after parturition. Removal of the pups of normal rats prevented the increases in activities of acetyl-CoA carboxylase and ATP citrate-lyase, but not of fatty acid synthetase, and administration of prolactin corticosterone or insulin did not stimulate activity. Tissue from suckled glands in which the ducts had been ligated at parturition showed no increase in the activities of acetyl-CoA carboxylase and ATP citrate-lyase within 24 h, whereas fatty acid synthetase activity was similar to that in the sham-operated contralateral glands. Foetoplacentectomy on day 18 increased the activity of fatty acid synthetase but not of acetyl-CoA carboxylase and ATP citrate-lyase; suckling of these dams by foster pups increased both acetyl-CoA carboxylase and ATP citrate-lyase.
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PMID:Initiation of lipogenic enzyme activities in rat mammary glands. 611 81

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

1. The effect of preincubation of extracts of lactating rat mammary gland with ATP, Mg2+ and micromolar concentrations of Ca2+ on the activity of acetyl-CoA carboxylase was studied. 2. Both Mg2+ and Ca2+ activated the enzyme. Activation with Mg2+ (5 mM) was larger than that with Ca2+ (calculated free Ca2+ concentration = 20-50 microM), but the activity decreased after reaching a peak. The activation obtained with Ca2+ was stable for up to 180 min. 3. Incubation with Ca2+ and Mg2+ together resulted in an activation that was slightly higher than that with Mg2+ only and was stable (compare the results for Ca2+ alone). 4. Preincubation in the absence of Mg2+, but not in the absence of Ca2+, resulted in the impairment of subsequent activation with either Mg2+ (when preincubation was with Ca2+ alone) or Mg2+ plus Ca2+. 5. KF (50 mM) prevented the activation of acetyl-CoA carboxylase by Ca2+ and Mg2+. 6. MgATP2- reversed (Mg2+ + Ca2+)-mediated activation and decreased the activity of acetyl-CoA carboxylase to about 10% of initial activity. Inhibition by ATP was unaffected by addition of cyclic AMP or cyclic AMP-dependent protein kinase inhibitor. 7. 32P was incorporated into acetyl-CoA carboxylase when incubations were carried out in the presence of [gamma-32P]ATP. Subsequent removal of ATP from the incubation medium resulted in rapid loss of 32P from acetyl-CoA carboxylase. 8. It is suggested that extracts of rat mammary gland contain endogenous protein kinase and phosphatase activities that modulate acetyl-CoA carboxylase activity through reversible phosphorylation and dephosphorylation. The phosphatase activity is sensitive to both Mg2+ and micromolar concentrations of Ca2+, whereas the kinase does not appear to be cyclic AMP-dependent.
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PMID:Regulation of acetyl-CoA carboxylase in rat mammary gland. Effects of incubation with Ca2+, Mg2+ and ATP on enzyme activity in tissue extracts. 612 11


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