Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:6.4.1.2 (acetyl-CoA carboxylase)
 2,876 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The formation of palmitoylcarnitine is catalyzed by carnitine palmitoyl-transferase (CPT-I) and this catalysis is the first committed step in beta-oxidation. The malonyl-CoA-inhibited isoform appears to be distinct from latent (CPT-II) activity, which is localized to the matrix side of the mitochondrial inner membrane. Sarcoplasmic reticulum from canine cardiac muscle was fractionated on a discontinuous sucrose density gradient into three major bands, all of which contained Ca(2+)-ATPase activity. Only the fraction that banded at a concentration of 38% surcrose was slightly contaminated by mitochondria. Peroxisomal uricase was low or absent in fractionated SR. All sarcoplasmic reticulum fractions contained malonyl-CoA-sensitive medium- (COT) and long-chain (CPT) carnitine acyltransferase activities. CPT activity decreased in sarcoplasmic reticulum when Triton X-100 was present. Carnitine acyltransferase activities were inactivated by preincubating the sarcoplasmic reticulum with the sulfhydryl reagent, 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB). In contrast, mitochondrial CPT-II activity was stable in the presence of DTNB and activated by Triton X-100. Western blots of mitochondria and sarcoplasmic reticulum fractions showed that the mitochondrial fractions reacted with antibody to mitochondrial CPT-II but not with SR protein when both were added at comparable specific activities. The data suggest that cardiac SR contains a unique malonyl-CoA-sensitive isoform of CPT, and that synthesis of acylcarnitine may occur in the microenvironment of Ca2+ transport, where the extent of production of acylcarnitine is controlled by cardiac acetyl-CoA carboxylase activity.
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PMID:Evidence for malonyl-CoA-sensitive carnitine acyl-CoA transferase activity in sarcoplasmic reticulum of canine heart. 162 48

Acetyl-coenzyme A carboxylase has been purified from the plastids of developing castor oil seeds. High concentrations of the enzyme are required for stability as well as the presence of dithiothreitol, glycerol, bicarbonate, Triton X-100, and polyvinyl-pyrrolidone. It has a molecular weight of approximately 528,000 and appears to be membrane associated. Acetyl-CoA carboxylase is active over a wide pH range with an optimum at 8.0. Arrhenius plots are biphasic. The enzyme displays normal Michaelis-Menten kinetics with limiting Michaelis constants of KATP, 0.1 mM; KHCO-3, 3.0 mM; and Kacetyl-CoA, 0.05 mM. Monovalent cations, such as K+ and Cs+, exert a small activating effect on the enzyme while a divalent cation, Mn2+ or Mg2+, is essential for activity. The enzyme does not appear to be highly regulated by cellular metabolites.
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PMID:Acetyl-coenzyme A carboxylase from the developing endosperm of Ricinus communis. I. Isolation and characterization. 613 95

The activity of acetyl-CoA carboxylase, measured in various ways, was studied in 15000g extracts of rat liver hepatocytes and compared with the rate of fatty acid synthesis in intact hepatocytes incubated with insulin or glucagon. Hepatocyte extracts were prepared by disruption of cells with a Dounce homogenizer or by solubilization with 1.5% (v/v) Triton X-100. Sucrose-density-gradient centrifugation demonstrated that the sedimentation coefficient of acetyl-CoA carboxylase from cell extracts was 30-35S, regardless of the conditions of incubation or disruption of hepatocytes. Solubilization of cells with 1.5% Triton X-100 yielded twice as much enzyme activity (measured by [14C]bicarbonate fixation) in the sucrose-gradient fractions as did cell disruption by the Dounce homogenizer. Analysis by high-performance liquid chromatography of acetyl-CoA carboxylase reaction mixtures showed that [14C]malonyl-CoA accounted for 10-60% of the total acid-stable radioactivity, depending on the method for disrupting hepatocytes and on the preincubation of the 15000g extract, with or without citrate, before assay. Under conditions in which incubation of cells with insulin or glucagon caused an activation or inhibition, respectively, of acetyl-CoA carboxylase, only 25% of the acid-stable radioactivity was [14C]malonyl-CoA and enzyme activity was only 13% (control), 16% (insulin), and 57% (glucagon) of the rate of fatty acid synthesis. Under conditions when up to 60% of the acid-stable radioactivity was [14C]malonyl-CoA and acetyl-CoA carboxylase activity was comparable with the rate of fatty acid synthesis, there was no effect of insulin or glucagon on enzyme activity.
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PMID:Studies on the assay, activity and sedimentation behaviour of acetyl-CoA carboxylase from isolated hepatocytes incubated with insulin or glucagon. 614 77

Acetyl-CoA carboxylase was purified 300-fold from rat liver, in the absence of added citrate, by precipitation from an 18,000g supernatant in the presence of Triton X-100 at 105,000g and 20 degrees C, followed by chromatography on phosphocellulose. Acetyl-CoA carboxylase activity in this preparation was activated by preincubation with GTP (0.1-2.0 mM) and with citrate (20 mM). Colchicine (10(-6)-10(-3) M) inhibited enzyme activity and counteracted the effects of GTP and citrate. Sucrose density gradient centrifugation demonstrated that GTP and citrate preincubation promoted the formation of the polymeric, active enzyme, while colchicine engendered disassembly. Preincubation of the purified acetyl-CoA carboxylase at 4 degrees C caused inactivation and disassembly, which was countered by preincubation at 37 degrees C in the presence of GTP or citrate. These results suggest that GTP, like citrate, activates acetyl-CoA carboxylase by enhancing the conversion of the protomeric form of the enzyme to its more active, polymeric state.
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PMID:Guanosine triphosphate and colchicine affect the activity and the polymeric state of acetyl-CoA carboxylase. 614 16

Concentrations of total CoAs in chloroplasts freshly isolated from spinach and peas were 10-20 microM, assuming a stromal volume of 66 microl per mg of chlorophyll. Acetyl-CoA and CoASH constituted at least 90% of the total CoA in freshly isolated chloroplasts. For a given chloroplast preparation, the concentration of endogenous acetyl-CoA was the same when extractions were performed using HClO4, trichloroacetic acid, propan-2-ol or chloroform/methanol, and the extracts analysed by quantitative HPLC after minimal processing. During fatty acid synthesis from acetate, concentrations of CoASH within spinach and pea chloroplasts varied from less than 0.1 to 5.0 microM. Malonyl-CoA concentrations were also very low (<0.1-3.0 microM) during fatty acid synthesis but could be calculated from radioactivity incorporated from [1-14C]acetate. Concentrations of CoASH in chloroplasts synthesizing fatty acids could be doubled in the presence of Triton X-100, suggesting that the detergent stimulates fatty acid synthesis by increasing the turnover rate of acyl-CoA. However, although taken up, exogenous CoASH (1 microM) did not stimulate fatty acid synthesis by permeabilized spinach chloroplasts. Calculated rates for acetyl-CoA synthetase, acetyl-CoA carboxylase and malonyl-CoA-acyl-carrier protein transacylase reactions at the concentrations of metabolites measured here are < 0.1-4% of the observed rates of fatty acid synthesis from acetate by isolated chloroplasts. The results suggest that CoA and its esters are probably confined within, and channelled through, the initial stages of a fatty acid synthase multienzyme complex.
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PMID:Stromal concentrations of coenzyme A and its esters are insufficient to account for rates of chloroplast fatty acid synthesis: evidence for substrate channelling within the chloroplast fatty acid synthase. 935 62

The committed step for de novo fatty acid biosynthesis is the carboxylation of acetyl-CoA catalyzed by acetyl-CoA carboxylase (ACCase). Plastidial ACCase from most plants is a multisubunit complex composed of multiple copies of four different polypeptides, biotin carboxyl carrier protein (BCCP), biotin carboxylase (BC), and carboxyltransferase (alpha-CT and beta-CT). Immunoblot analyses revealed these four proteins were mostly (69% of total) associated with a 17,000 g insoluble fraction from lysed pea chloroplasts. Under the same conditions only 8% of ribulose-1,5-bisphosphate carboxylase was associated with this insoluble fraction. BCCP and biotin carboxylase BC subunits freely dissociated from 17 kg insoluble fractions under high ionic strength conditions, whereas alpha-CT and beta-CT subunits remained tightly associated. Both CT subunits were highly enriched in envelope versus stroma and thylakoid preparations whereas BC and BCCP subunits were predominantly stromal-localized due to partial dissociation. Rapid solubilization of intact chloroplasts with Triton X-100 followed by centrifugation at 30 kg resulted in a pellet that was up to 8-fold enriched in ACCase activity and 21-fold enriched in BC activity. Triton-insoluble 30 kg pellets were reduced in lipid and chlorophyll content but enriched in chloroplast DNA due to the isolation of nucleoid particles. However, ACCase was not directly associated with nucleoids since enzymatic digestion of DNA or RNA had no effect on the association with Triton-insoluble matter. The amount of Triton-insoluble ACCase was similar in chloroplasts isolated from dark- or light-adapted leaves suggesting transitory starch granules were also not involved in this association. It is proposed that ACCase is associated with envelope membranes through interactions with an unidentified integral membrane protein.
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PMID:The multisubunit acetyl-CoA carboxylase is strongly associated with the chloroplast envelope through non-ionic interactions to the carboxyltransferase subunits. 1205 35

Proteomic analysis of a Triton X-100 insoluble, 30,000 x g pellet from purified pea chloroplasts resulted in the identification of 179 nonredundant proteins. This chloroplast fraction was mostly depleted of chloroplast membranes since only 23% and 9% of the identified proteins were also observed in envelope and thylakoid membranes, respectively. One of the most abundant proteins in this fraction was sulfite reductase, a dual function protein previously shown to act as a plastid DNA condensing protein. Approximately 35 other proteins known (or predicted) to be associated with high-density protein-nucleic acid particles (nucleoids) were also identified including a family of DNA gyrases, as well as proteins involved in plastid transcription and translation. Although nucleoids appeared to be the predominant component of 30k x g Triton-insoluble chloroplast preparations, multi-enzyme protein complexes were also present including each subunit to the pyruvate dehydrogenase and acetyl-CoA carboxylase multi-enzyme complexes, as well as a proposed assembly of the first three enzymes of the Calvin cycle. Approximately 18% of the proteins identified were annonated as unknown or hypothetical proteins and another 20% contained "putative" or "like" in the identifier tag. This is the first proteomic characterization of a membrane-depleted, high-density fraction from plastids and demonstrates the utility of this simple procedure to isolate intact macromolecular structures from purified organelles for analysis of protein-protein and protein-nucleic acid interactions.
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PMID:Proteomic characterization of a triton-insoluble fraction from chloroplasts defines a novel group of proteins associated with macromolecular structures. 1582 27

The pyruvate dehydrogenase complex from pea (Pisum sativum L.) mitochondria was purified 23-fold by high speed centrifugation and glycerol gradient fractionation. The complex had a s(20,w) of 47.5S but this is a minimal value since the complex is unstable. The complex is specific for NAD(+) and pyruvate; NADP(+) and other keto acids give no reaction. Mg(2+), thiamine pyrophosphate, and cysteine are also required for maximal activity. The pH optimum for the complex was between 6.5 and 7.5.Continuous sucrose density gradients were used to separate castor bean (Ricinus communis L.) endosperm proplastids from mitochondria. Pyruvate dehydrogenase complex activity was found to be coincident with the proplastid peak on all of the gradients. Some separation of proplastids and mitochondria could be achieved by differential centrifugation and the ratios of the activities of the pyruvate dehydrogenase complex to succinic dehydrogenase and acetyl-CoA carboxylase to succinic dehydrogenase were consistent with both the pyruvate dehydrogenase complex and acetyl-CoA carboxylase being present in the proplastid. The proplastid fraction has to be treated with a detergent, Triton X-100, before maximal activity of the pyruvate dehydrogenase complex activity is expressed, indicating that it is bound in the organelle. The complex had a sharp pH optimum of 7.5. The complex required added Mg(2+), cysteine, and thiamine pyrophosphate for maximal activity but thiamine pyrophosphate was inhibitory at higher concentrations.
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PMID:Pyruvate dehydrogenase complex from higher plant mitochondria and proplastids. 1665 53

The mammary epithelium produces numerous lipid droplets during lactation and secretes them in plasma membrane-enclosed vesicles known as milk fat globules. The biogenesis of such fat globules is considered to provide a model for clarifying the mechanisms of lipogenesis in mammals. In the present study, we identified acetyl coenzyme A carboxylase, ATP citrate lyase, and fatty acid synthase in mouse milk. Fractionation of milk showed that these three enzymes were located predominantly in milk fat globules. The three enzymes were resistant to trypsin digestion without Triton X-100, indicating that they were not located on the outer surface of the globules and thus associated with the precursors of the globules before secretion. When a low dose of rapamycin, an inhibitor of the mammalian target of rapamycin (mTOR), was injected into lactating mice, the levels of the three enzymes in milk were decreased within 3h after injection. Since the protein levels of the three enzymes in tissues were not obviously altered by this short-term treatment, known transcriptional control by mTOR signaling was unlikely to account for this decrease in their levels in milk. Our findings suggest a new, putatively mTOR-dependent localization of the three enzymes for de novo lipogenesis.
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PMID:Secretion of three enzymes for fatty acid synthesis into mouse milk in association with fat globules, and rapid decrease of the secreted enzymes by treatment with rapamycin. 2128 98