Polymer-protomer transition of acetyl-CoA carboxylase occurs in vivo and varies with nutritional conditions.

The ‘initial’ (I), endogenous phosphatase-activated (A) and citrate-activated (C) activities of acetyl-CoA carboxylase were measured in mammary-gland extracts of pregnant and lactating rats. There was a 10-fold increase in the A and C enzyme activities in the transition from early to peak lactation [cf. data of Mackall & Lane (1977) Biochem. J.162, 635–642], but there was no significant increase in the ratio of the initial activity to the A and C activities of the enzyme. Starvation (24h) or short-term (3h) streptozotocin-induced diabetes both resulted in a 40% decrease in I/A and I/C activity ratios. In starvation this was accompanied by a decrease in the absolute values of the A and C activities such that the initial activity in mammary glands of starved animals was 45% that in glands from fed animals. Insulin treatment of starved or diabetic animals 60min before killing increased the I activity without affecting the A or C enzyme activities. Removal of the pups for 24h from animals in peak lactation (weaning) resulted in a marked but similar decrease in all three activities such that, although the initial activity was only 10% of that in suckled animals, the I/A and I/C activity ratios remained high and unaltered. Inhibition of prolactin secretion by injection of 2-bromo-α-ergocryptine gave qualitatively similar results to those during weaning. Simultaneous administration of ovine prolactin completely prevented the effects of bromoergocryptine. It is suggested that the initial activity of acetyl-CoA carboxylase in rat mammary gland is regulated by at least two parallel mechanisms: (i) an acute regulation of the proportion of the enzyme in the active state and (ii) a longer-term modulation of enzyme concentration in the gland. Insulin appeared to mediate its acute effects through mechanism (i), whereas prolactin had longer-term effects on enzyme concentration in the gland. A comparison of initial enzyme activities (I) obtained in the present study with rates of lipogenesis measured in vivo [Agius & Williamson (1980) Biochem. J.192, 361–364; Munday & Williamson (1981) Biochem. J.196, 831–837] gave good agreement between the two sets of data for all conditions studied except for 24h-starved and streptozotocin-diabetic animals. It is suggested that acetyl-CoA carboxylase activity is rate-limiting for lipogenesis in the mammary gland in normal, fed, suckled or weaned animals but that in starved and short-term diabetic animals changes in the activity of the enzyme by covalent modification alone may not be sufficient to maintain the enzyme in its rate-limiting role.

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P5375A default in acetyl-CoA carboxylase phosphorylation increases thrombus growth in vitro and in vivo, and in a collagen-dependent manner

European Heart Journal ◽

10.1093/eurheartj/ehx493.p5375 ◽

2017 ◽

Vol 38 (suppl_1) ◽

Author(s):

S. Lepropre ◽

S. Kautbally ◽

L. Bertrand ◽

G.R. Steinberg ◽

B.E. Kemp ◽

...

Keyword(s):

Dependent Manner ◽

Acetyl Coa Carboxylase ◽

Acetyl Coa

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Kinetic studies on two isoforms of acetyl-CoA carboxylase from maize leaves

Biochemical Journal ◽

10.1042/bj3180997 ◽

1996 ◽

Vol 318 (3) ◽

pp. 997-1006 ◽

Cited By ~ 36

Author(s):

Derek HERBERT ◽

Lindsey J PRICE ◽

Claude ALBAN ◽

Laure DEHAYE ◽

Dominique JOB ◽

...

Keyword(s):

Kinetic Studies ◽

Product Inhibition ◽

Hill Equation ◽

Acetyl Coa Carboxylase ◽

Acetyl Coa ◽

Maize Leaves ◽

Ic50 Values ◽

A Minor ◽

The Hill

The steady-state kinetics of two multifunctional isoforms of acetyl-CoA carboxylase (ACCase) from maize leaves (a major isoform, ACCase1 and a minor isoform, ACCase2) have been investigated with respect to reaction mechanism, inhibition by two graminicides of the aryloxyphenoxypropionate class (quizalofop and fluazifop) and some cellular metabolites. Substrate interaction and product inhibition patterns indicated that ADP and Pi products from the first partial reaction were not released before acetyl-CoA bound to the enzymes. Product inhibition patterns did not match exactly those predicted for an ordered Ter Ter or a random Ter Ter mechanism, but were close to those postulated for an ordered mechanism. ACCase2 was about 1/2000 as sensitive as ACCase1 to quizalofop but only about 1/150 as sensitive to fluazifop. Fitting inhibition data to the Hill equation indicated that binding of quizalofop or fluazifop to ACCase1 was non-cooperative, as shown by the Hill constant (napp) values of 0.86 and 1.16 for quizalofop and fluazifop respectively. Apparent inhibition constant values (K´ from the Hill equation) for ACCase1 were 0.054 µM for quizalofop and 21.8 µM for fluazifop. On the other hand, binding of quizalofop or fluazifop to ACCase2 exhibited positive co-operativity, as shown by the napp values of 1.85 and 1.59 for quizalofop and fluazifop respectively. K´ values for ACCase2 were 1.7 mM for quizalofop and 140 mM for fluazifop. Kinetic parameters for the co-operative binding of quizalofop to maize ACCase2 were close to those of another multifunctional ACCase of limited sensitivity to graminicide, ACC220 from pea. Inhibition of ACCase1 by quizalofop was mixed-type with respect to acetyl-CoA or ATP, but the concentration of acetyl-CoA had the greater effect on the level of inhibition. Neither ACCase1 nor ACCase2 was appreciably sensitive to CoA esters of palmitic acid (16:0) or oleic acid (18:1). Approximate IC50 values were 10 µM (ACCase2) and 50 µM (ACCase1) for both CoA esters. Citrate concentrations up to 1 mM had no effect on ACCase1 activity. Above this concentration, citrate was inhibitory. ACCase2 activity was slightly stimulated by citrate over a broad concentration range (0.25–10 mM). The significance of possible effects of acyl-CoAs or citrate in vivo is discussed.

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RNA-seq reveals novel mechanistic targets of withaferin A in prostate cancer cells

Carcinogenesis ◽

10.1093/carcin/bgaa009 ◽

2020 ◽

Vol 41 (6) ◽

pp. 778-789 ◽

Cited By ~ 5

Author(s):

Su-Hyeong Kim ◽

Eun-Ryeong Hahm ◽

Krishna B Singh ◽

Sruti Shiva ◽

Jacob Stewart-Ornstein ◽

...

Keyword(s):

Prostate Cancer ◽

Fatty Acid ◽

Fatty Acid Synthase ◽

Acid Synthesis ◽

Acetyl Coa Carboxylase ◽

Rna Seq ◽

Atp Citrate Lyase ◽

Acetyl Coa ◽

Citrate Lyase

Abstract Withaferin A (WA) is a promising phytochemical exhibiting in vitro and in vivo anticancer activities against prostate and other cancers, but the mechanism of its action is not fully understood. In this study, we performed RNA-seq analysis using 22Rv1 human prostate cancer cell line to identify mechanistic targets of WA. Kyoto Encyclopedia of Genes and Genomes pathway analysis of the differentially expressed genes showed most significant enrichment of genes associated with metabolism. These results were validated using LNCaP and 22Rv1 human prostate cancer cells and Hi-Myc transgenic mice as models. The intracellular levels of acetyl-CoA, total free fatty acids and neutral lipids were decreased significantly following WA treatment in both cells, which was accompanied by downregulation of mRNA (confirmed by quantitative reverse transcription-polymerase chain reaction) and protein levels of key fatty acid synthesis enzymes, including ATP citrate lyase, acetyl-CoA carboxylase 1, fatty acid synthase and carnitine palmitoyltransferase 1A. Ectopic expression of c-Myc, but not constitutively active Akt, conferred a marked protection against WA-mediated suppression of acetyl-CoA carboxylase 1 and fatty acid synthase protein expression, and clonogenic cell survival. WA was a superior inhibitor of cell proliferation and fatty acid synthesis in comparison with known modulators of fatty acid metabolism including cerulenin and etomoxir. Intraperitoneal WA administration to Hi-Myc transgenic mice (0.1 mg/mouse, three times/week for 5 weeks) also resulted in a significant decrease in circulating levels of total free fatty acids and phospholipids, and expression of ATP citrate lyase, acetyl-CoA carboxylase 1, fatty acid synthase and carnitine palmitoyltransferase 1A proteins in the prostate in vivo.

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Expression, biotinylation and purification of a biotin-domain peptide from the biotin carboxy carrier protein of Escherichia coli acetyl-CoA carboxylase

Biochemical Journal ◽

10.1042/bj3020881 ◽

1994 ◽

Vol 302 (3) ◽

pp. 881-887 ◽

Cited By ~ 97

Author(s):

A Chapman-Smith ◽

D L Turner ◽

J E Cronan ◽

T W Morris ◽

J C Wallace

Keyword(s):

Escherichia Coli ◽

Exchange Chromatography ◽

Carrier Protein ◽

Acetyl Coa Carboxylase ◽

Acetyl Coa ◽

E Coli ◽

Biotin Ligase ◽

Domain Peptide

A protein segment consisting of the C-terminal 87 residues of the biotin carboxy carrier protein from Escherichia coli acetyl-CoA carboxylase was overexpressed in E. coli. The expressed biotin-domain peptide can be fully biotinylated by coexpression with a plasmid that overproduces E. coli biotin ligase. The extent of biotinylation was limited in vivo, but could be taken to completion in cell lysates on addition of ATP and biotin. We used the coexpression of biotin ligase and acceptor protein to label the biotin-domain peptide in vitro with [3H]biotin, which greatly facilitated development of a purification procedure. The apo (unbiotinylated) form of the protein was prepared by induction of biotin-domain expression in a strain lacking the biotin-ligase-overproduction plasmid. The apo domain could be separated from the biotinylated protein by ion-exchange chromatography or non-denaturing PAGE, and was converted into the biotinylated form of the peptide on addition of purified biotin ligase. The identify of the purified biotin-domain peptide was confirmed by N-terminal sequence analysis, amino acid analysis and m.s. The domain was readily produced and purified in sufficient quantities for n.m.r. structural analysis.

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Quantitation by immunoblotting of the in vivo induction and subcellular distribution of hepatic acetyl-CoA carboxylase

Archives of Biochemistry and Biophysics ◽

10.1016/0003-9861(88)90575-9 ◽

1988 ◽

Vol 264 (1) ◽

pp. 103-113 ◽

Cited By ~ 5

Author(s):

Joseph L. Evans ◽

Lee A. Witters

Keyword(s):

Subcellular Distribution ◽

Acetyl Coa Carboxylase ◽

Acetyl Coa ◽

In Vivo Induction

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Insulin activation of lipogenesis in isolated mammary acini from lactating rats fed on a high-fat diet. Evidence that acetyl-CoA carboxylase is a site of action

Biochemical Journal ◽

10.1042/bj2420905 ◽

1987 ◽

Vol 242 (3) ◽

pp. 905-911 ◽

Cited By ~ 16

Author(s):

M R Munday ◽

D H Williamson

Keyword(s):

Glucose Uptake ◽

High Fat Diet ◽

Acetyl Coa Carboxylase ◽

High Fat ◽

Acetyl Coa ◽

Carboxylase Activity ◽

Glucose Incorporation ◽

Site Of Action ◽

A Site

Feeding lactating rats on high-fat cheese crackers in addition to laboratory chow increased the dietary intake of fat from 2 to 20% of the total weight of food eaten and decreased mammary-gland lipogenesis in vivo by approx. 50%. This lipogenic inhibition was also observed in isolated mammary acini, where it was accompanied by decreased glucose uptake. These inhibitions were completely reversed by incubation with insulin. Insulin had no effect on the rate of glucose transport into acini, nor on pyruvate dehydrogenase activity as estimated by the accumulation of pyruvate and lactate, suggesting that these are not the sites of lipogenic inhibition. Insulin stimulated the incorporation of [1-14C]acetate into lipid in acini from high-fat-fed rats. In the presence of alpha-cyanohydroxycinnamate, a potent inhibitor of mitochondrial pyruvate transport, and with glucose as the sole substrate, neither [1-14C]glucose incorporation into lipid nor glucose uptake were stimulated by insulin. Insulin did stimulate the incorporation of [1-14C]acetate into lipid in the presence of alpha-cyanohydroxycinnamate, and this was accompanied by an increase in glucose uptake by the acini. This indicated that increased glucose uptake was secondary to the stimulation of lipogenesis by insulin, which therefore must occur via activation of a step in the pathway distal to mitochondrial pyruvate transport. Insulin stimulated acetyl-CoA carboxylase activity measured in crude extracts of acini from high-fat-fed rats, restoring it to values close to those of chow-fed controls. The effects of insulin on acetyl-CoA carboxylase activity and lipogenesis were not antagonized by adrenaline or dibutyryl cyclic AMP.

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