Partial purification of rat liver cytoplasmic acetyl-CoA synthetase; Characterization of some properties

Imesch, Elisabeth; Rous, Simonne

doi:10.1016/0020-711x(84)90146-0

Cited by 15 publications

(10 citation statements)

References 25 publications

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“…The apparent K m values for acetate, CoA, and ATP were 73, 11, and 245 M, respectively, as calculated from Lineweaver-Burke plots. These values are similar to the published apparent K m values for ACS that was partially purified from rat liver (15) except that the apparent affinity for CoA was 4-fold higher for the recombinant enzyme.…”

Section: Figsupporting

confidence: 79%

“…The hamster cDNA fragment was used to isolate the full-length human ACS cDNA, and this was used to produce active ACS enzyme by transfection into human cells in tissue culture. The recombinant enzyme functioned as a monomer of 80 kDa whose kinetic properties were similar to the previously described rat liver ACS (15). Using the human cDNA as a probe, we demonstrated that ACS mRNA is increased in tissue culture cells when nSREBPs enter the nucleus in response to cholesterol deprivation, and this increase is prevented when SREBP activation is blocked with sterols.…”

Section: Figmentioning

confidence: 76%

“…The reaction requires ATP and proceeds through the formation of an acetyl adenylate (12,13). ACS activity has been studied biochemically in animal cells (14), and the enzyme has been partially purified from rat liver (15), but the gene has not been cloned, and the molecular structure of the animal cell enzyme is unknown. Here, we provide the first molecular characterization of animal ACS, and we demonstrate that its mRNA is controlled by SREBPs.…”

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confidence: 99%

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Molecular Characterization of Human Acetyl-CoA Synthetase, an Enzyme Regulated by Sterol Regulatory Element-binding Proteins

Luong¹,

Hannah²,

Brown³

et al. 2000

Journal of Biological Chemistry

231

204

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Through suppressive subtractive hybridization, we identified a new gene whose transcription is induced by sterol regulatory element-binding proteins (SREBPs). The gene encodes acetyl-CoA synthetase (ACS), the cytosolic enzyme that activates acetate so that it can be used for lipid synthesis or for energy generation. ACS genes were isolated previously from yeast, but not from animal cells. Recombinant human ACS was produced by expressing the cloned cDNA transiently in human cells. After purification by nickel chromatography, the 701-amino acid cytosolic enzyme was shown to function as a monomer. The recombinant enzyme produced acetylCoA from acetate in a reaction that required ATP. As expected for a gene controlled by SREBPs, ACS mRNA was induced when cultured cells were deprived of sterols and repressed by sterol addition. The pattern of regulation resembled the regulation of enzymes of fatty acid synthesis. ACS mRNA was also elevated in livers of transgenic mice that express dominant-positive versions of all three isoforms of SREBP. We conclude that ACS mRNA, and hence the ability of cells to activate acetate, is regulated by SREBPs in parallel with fatty acid synthesis in animal cells.Sterol regulatory element-binding proteins (SREBPs) 1 are transcription factors that activate more than 20 genes that produce enzymes required for the synthesis of cholesterol and unsaturated fatty acids in animal cells (1-3). The SREBPs differ from other transcription factors because they are synthesized as membrane-bound proteins whose active fragments must be released by proteolysis in order to enter the nucleus and activate transcription. The proteolytic release of the nuclear fragments is controlled by the cholesterol content of the cell; release is rapid when cells are depleted of cholesterol, and it is blocked when cholesterol overaccumulates (1).In tissue culture cells and in livers of transgenic mice, nuclear SREBPs (nSREBPs) increase the levels of mRNAs encoding multiple enzymes in the cholesterol biosynthetic pathway, including 3-hydroxy-3-methylglutaryl coenzyme A synthase (HMG-CoA synthase), HMG-CoA reductase, farnesyl diphosphate synthase, squalene synthase, lanosterol demethylase, and others. In the fatty acid biosynthetic pathway, SREBPs increase the mRNAs encoding acetyl-CoA carboxylase, fatty acid synthetase, and stearoyl-CoA desaturase. As a result of these changes, there is a massive increase in the content of unsaturated fatty acids and cholesterol in livers of transgenic mice that overexpress either of two nuclear isoforms of SREBP (nSREBP-1a and nSREBP-2) (4, 5).In addition to increasing enzymes that participate directly in lipid biosynthesis, the SREBPs increase the mRNA encoding ATP-citrate lyase, which is the major source of the acetyl-CoA that is the ultimate building block for lipid synthesis (6, 7). The SREBPs also increase the mRNAs for three enzymes that supply the NADPH that is needed for lipogenesis (malic enzyme, glucose-6-phosphate dehydrogenase, and 6-phosphogluconate dehydrogenase) (8).In th...

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Section: Figsupporting

confidence: 79%

Section: Figmentioning

confidence: 76%

mentioning

confidence: 99%

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Molecular Characterization of Human Acetyl-CoA Synthetase, an Enzyme Regulated by Sterol Regulatory Element-binding Proteins

Luong¹,

Hannah²,

Brown³

et al. 2000

Journal of Biological Chemistry

231

204

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“…The enzyme has also been purified from mammals (17) and from several species of fungus (6,28,32), and these have the same substrate specificity as the prokaryotic enzymes. Hence, the discovery of an ADP-dependent ACS in cell extracts of the hyperthermophilic, acetate-producing archaeon P. furiosus by Schäfer and Schönheit (37) was of some significance.…”

Section: Discussionmentioning

confidence: 99%

“…ACS has been purified from several organisms able to utilize acetate, including bacteria (32), methanogenic archaea (18), and eukaryotes (6,17,28). All are homodimeric enzymes with a subunit M r of about 70,000.…”

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confidence: 99%

Purification and characterization of two reversible and ADP-dependent acetyl coenzyme A synthetases from the hyperthermophilic archaeon Pyrococcus furiosus

1996

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Pyrococcus furiosus is a strictly anaerobic archaeon (archaebacterium) that grows at temperatures up to 105؇C by fermenting carbohydrates and peptides. Cell extracts have been previously shown to contain an unusual acetyl coenzyme A (acetyl-CoA) synthetase (ACS) which catalyzes the formation of acetate and ATP from acetyl-CoA by using ADP and phosphate rather than AMP and PP i . We show here that P. furiosus contains two distinct isoenzymes of ACS, and both have been purified.

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Organic Synthesis with Ligases

2022

Enzyme‐Based Organic Synthesis

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Partial purification of rat liver cytoplasmic acetyl-CoA synthetase; Characterization of some properties

Cited by 15 publications

References 25 publications

Molecular Characterization of Human Acetyl-CoA Synthetase, an Enzyme Regulated by Sterol Regulatory Element-binding Proteins

Molecular Characterization of Human Acetyl-CoA Synthetase, an Enzyme Regulated by Sterol Regulatory Element-binding Proteins

Purification and characterization of two reversible and ADP-dependent acetyl coenzyme A synthetases from the hyperthermophilic archaeon Pyrococcus furiosus

Organic Synthesis with Ligases

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