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1.
Polymer-protomer transition of acetyl-CoA carboxylase occurs in vivo and varies with nutritional conditions 总被引:2,自引:0,他引:2
B A Ashcraft W S Fillers S L Augustine S D Clarke 《The Journal of biological chemistry》1980,255(21):10033-10035
The protomeric form of purified acetyl coenzyme A carboxylase is inactivated by the binding of avidin to the biotinyl prosthetic group; the catalytically active filamentous form of the enzyme is resistant to avidin. This differential sensitivity to avidin was used to examine the influence of nutritional state on the proportion of polymeric and protomeric carboxylase occurring in avian liver. Hepatic carboxylase was 80% avidin-resistant (polymeric) in the fed chick. Food deprivation for 2 and 6 h reduced the avidin resistance to 54% and 30%, respectively. Similarly, within 1 h after fat intubation, the fraction of polymeric carboxylase had significantly decreased. Accompanying the change in carboxylase transformation was a comparable reduction in 3H2O incorporation into liver fatty acid. These data indicate that the protomer-polymer transition defined for purified acetyl-CoA carboxylase also occurs with the enzyme in vivo and that a lower polymer/protomer ratio is associated with reduced rates of fatty acid synthesis. 相似文献
2.
Fatty acid-mediated disaggregation of acetyl-CoA carboxylase in isolated liver cells 总被引:1,自引:0,他引:1
In recent years the rapid regulation of acetyl-CoA (AcCoA) carboxylase (EC 6.4.1.2) has become of major interest because of the important role of malonyl-CoA in fatty acid synthesis, ketogenesis, and triglyceride production. AcCoA carboxylase is acutely regulated by two mechanisms: 1) phosphorylation-dephosphorylation and 2) polymer-protomer transition. Until recently polymer-protomer transition of AcCoA carboxylase in vivo has escaped detection. We developed a technique that estimates the intracellular proportion of polymer and protomer forms of AcCoA carboxylase based on the differential sensitivity of polymeric and protomeric AcCoA carboxylase to avidin inactivation. When the enzyme is in its highly aggregated conformation, the biotin prosthetic group of AcCoA carboxylase is protected from avidin binding. Thus the polymeric AcCoA carboxylase is more resistant than the protomeric conformation to avidin inactivation. Utilizing this technique with isolated liver cells we have been able to develop a model for the involvement of free fatty acids and glucagon in regulating polymer-protomer transition of AcCoA carboxylase, and the role of polymer as an intracellular determinant of AcCoA carboxylase activity. Our data suggest that the physiological regulation of AcCoA carboxylase involves the interaction of the phosphorylation mechanism with fatty acid-induced depolymerization. We propose that during periods of food deprivation the elevation in fatty acid-CoA esters promotes depolymerization of AcCoA carboxylase. In addition, glucagon induces phosphorylation of AcCoA carboxylase, which inhibits the enzyme's activity and facilitates acyl-CoA binding and depolymerization. The two separate mechanisms for regulating hepatic AcCoA carboxylase may work in concert to modulate the level of the regulatory metabolite malonyl-CoA. 相似文献
3.
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 °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?3m) 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 °C caused inactivation and disassembly, which was countered by preincubation at 37 °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. 相似文献
4.
If acetyl-CoA carboxylase in epididymal fat tissue is subject to control by convalent modification as in the case of the liver enzyme, catalytically different forms of carboxylase should exist, independent of polymerization. By treating epididymal fat tissue in culture with epinephrine, we have demonstrated catalytically less active forms of acetyl-CoA carboxylase. The catalytically less active forms of the enzyme reacted to antibody with the same efficiency as the active form of carboxylase. However, the less active enzyme formed by epinephrine treatment of tissues has a sedimentation constant of 30 to 35 S, whereas that of the enzyme from control tissue is 45 S. Incubation of the less active forms of the carboxylase with 10 mM citrate and up to 10 mg/ml of bovine serum albumin activated the enzyme without any change in the sedimentation constant. Therefore, the less active forms of the carboxylase formed as a result of epinephrine treatment are not due to the depolymerization of polymeric forms (45 S) to the protomeric forms (17 to 20 S), but to the formation of intermediate species of carboxylase which cannot form polymeric enzyme (45 S) in the presence of high concentrations of citrate. 相似文献
5.
1. Acetyl-CoA carboxylase activity was measured in extracts of rat epididymal fat-pads either on preparation of the extracts (initial activity) or after incubation of the extracts with citrate (total activity). In the presence of glucose or fructose, brief exposure of pads to insulin increased the initial activity of acetyl-CoA carboxylase; no increase occurred in the absence of substrate. Adrenaline in the presence of glucose and insulin decreased the initial activity. None of these treatments led to a substantial change in the total activity of acetyl-CoA carboxylase. A large decrease in the initial activity of acetyl-CoA carboxylase also occurred with fat-pads obtained from rats that had been starved for 36h although the total activity was little changed by this treatment. 2. Conditions of high-speed centrifugation were found which appear to permit the separation of the polymeric and protomeric forms of the enzyme in fat-pad extracts. After the exposure of the fat-pads to insulin (in the presence of glucose), the proportion of the enzyme in the polymeric form was increased, whereas exposure to adrenaline (in the presence of glucose and insulin) led to a decrease in enzyme activity. 3. These changes are consistent with a role of citrate (as activator) or fatty acyl-CoA thioesters (as inhibitors) in the regulation of the enzyme by insulin and adrenaline; no evidence that the effects of these hormones involve phosphorylation or dephosphorylation of the enzyme could be found. 4. Changes in the whole tissue concentration of citrate and fatty acyl-CoA thioesters were compared with changes in the initial activity of acetyl-CoA carboxylase under a variety of conditions of incubation. No correlation between the citrate concentration and the initial enzyme activity was evident under any condition studied. Except in fat-pads which were exposed to insulin there was little inverse correlation between the concentration in the tissue of fatty acyl-CoA thioesters and the initial activity of acetyl-CoA carboxylase. 5. It is suggested that changes in the concentration of free fatty acyl-CoA thioesters (which may not be reflected in whole tissue concentrations of these metabolites) may be important in the regulation of the activity of acetyl-CoA carboxylase. The possibility is discussed that the concentration of free fatty acyl-CoA thioesters may be controlled by binding to a specific protein with properties similar to albumin. 相似文献
6.
Digitonin treatment of chick liver cells in monolayer culture perforates the plasma membrane, causing release of acetyl-CoA carboxylase and other cytosolic enzymes. The rate of carboxylase release is affected by conditions known to alter the position of the protomer-polymer (filament) equilibrium of the enzyme. Citrate, an allosteric activator of the carboxylase, induces polymerization of the protomeric avidin-sensitive form giving rise to the avidin-insensitive polymeric filamentous form. When cells are exposed to N6,O2-dibutyryl cyclic adenosine 3':5'-monophosphate which lowers intracellular citrate levels, the rate of carboxylase release from digitonin-treated cells is greatly accelerated. The presence of avidin, which rapidly enters the cell during digitonin treatment, inactivates carboxylase under conditions that promote depolymerization and rapid release, but not under conditions which promote polymerization and slow release. These findings indicate that carboxylase filaments exist in the intact chick liver cell when the cytoplasmic citrate level is high and undergo depolymerization when citrate levels fall. 相似文献
7.
Acetyl-CoA carboxylase catalyzes the first committed step in the synthesis of fatty acids. Because fatty acids are required during myelination in the developing brain, it was proposed that the level of acetyl-CoA carboxylase may be highest in embryonic brain. The presence of acetyl-CoA carboxylase activity was detected in chick embryo brain. Its activity varied with age, showing a peak in the 17-18-day-old embryo and decreasing thereafter. The enzyme, affinity-purified from 18-day-old chick embryo brain, appeared as a major protein band on polyacrylamide electrophoresis gels in the presence of sodium dodecyl sulfate (Mr 265,000), indistinguishable from the 265 kDa isozyme of liver acetyl-CoA carboxylase. It had significant activity (Sp act = 1.1 mumol/min per mg protein) in the absence of citrate. There was a maximum stimulation of only 25% in the presence of citrate. Dephosphorylation using [acetyl-CoA carboxylase] phosphatase 2 did not result in activation of the enzyme. Palmitoyl-CoA (0.1 mM) and malonyl-CoA (1 mM) inhibited the activity to 95% and 71%, respectively. Palmitoylcarnitine, however, did not show significant inhibition. The enzyme was inhibited (greater than 95%) by avidin; however, avidin did not show significant inhibition in the presence of excess biotin. The enzyme was also inhibited (greater than 90%) by antibodies against liver acetyl-CoA carboxylase. An immunoblot or avidin-blot detected only one protein band (Mr 265,000) in preparations from chick embryo brain or adult liver. These observations suggest that acetyl-CoA carboxylase is present in embryonic brain and that the enzyme appears to be similar to the 265 kDa isozyme of liver. 相似文献
8.
The polymerization of acetyl-CoA carboxylase 总被引:2,自引:0,他引:2
Citrate, an allosteric activator of acetyl-CoA carboxylase, induces polymerization of an inactive protomeric form of the enzyme into an active filamentous form composed of 10-20 protomers. The light-scattering properties of the carboxylase were used to study the kinetics of its polymerization and depolymerization. From stopped flow kinetic studies, we have established that polymerization is a second order process, with a second order rate constant of 597,000 M-1 s-1. There appear to be two steps which limit polymerization of the inactive carboxylase protomer: 1) a rapid citrate-induced conformational change which is independent of enzyme concentration and leads to an active protomeric form of the enzyme (Beaty, N. B., and Lane, M. D. (1983) J. Biol. Chem. 258, 13043-13050, preceding paper) and 2) the dimerization of the active protomer, which constitutes the first step of polymerization and is enzyme concentration-dependent. Dimerization is the rate-limiting step of acetyl-CoA carboxylase polymerization. Depolymerization of fully polymerized acetyl-CoA carboxylase is caused by malonyl-CoA, ATP X Mg, and Mg2+. Both malonyl-CoA and ATP X Mg (and HCO-3) compete with citrate in the maintenance of a given state of the protomer-polymer equilibrium apparently by carboxylating the enzyme to form enzyme-biotin-CO-2 which destablizes the polymeric form. Free citrate is the species responsible for polymerizing the enzyme and Mg2+ causes depolymerization of the enzyme by lowering the concentration of free citrate. 相似文献
9.
M R Lakshmanan C M Nepokroeff M Kim J W Porter 《Archives of biochemistry and biophysics》1975,169(2):737-745
Hepatocytes were isolated at specified times from livers of diabetic and insulin-treated diabetic rats during the course of a 48-h refeeding of a fat-free diet to previously fasted rats. The rates of synthesis of fatty acid synthetase and acetyl-CoA carboxylase in the isolated cells were determined as a function of time of refeeding by a 2-h incubation with l-[U-14C]leucine. Immunochemical methods were employed to determine the amount of radioactivity in the fatty acid synthetase and acetyl-CoA carboxylase proteins. The amount of radioactivity in the fatty acid synthetase synthesized by the isolated cells was also determined following enzyme purification of the enzyme to homogeneity. Enzyme activities of the fatty acid synthetase and acetyl-CoA carboxylase in the cells were measured by standard procedures. The results show that isolated liver cells obtained from insulintreated diabetic rats retain the capacity to synthesize fatty acid synthetase and acetyl-CoA carboxylase. The rate of synthesis of the fatty acid synthetase in the isolated cells was similar to the rate found in normal refed animals in in vivo experiments [Craig et al. (1972) Arch. Biochem. Biophys. 152, 619–630; Lakshmanan et al. (1972) Proc. Nat. Acad. Sci. USA69, 3516–3519]. In addition the relative rate of synthesis of fatty acid synthetase was stimulated greater than 20-fold in the diabetic animals treated with insulin. Immunochemical assays, when compared with enzyme activities, indicated the presence of an immunologically reactive, but enzymatically inactive, form or “apoenzyme” for both the fatty acid synthetase and acetyl-CoA carboxylase. The synthesis of these immunoreactive and enzymatically inactive species of protein, as well as the synthesis of the “holoenzyme” forms of both enzymes, requires insulin. 相似文献
10.
Quantitation by immunoblotting of the in vivo induction and subcellular distribution of hepatic acetyl-CoA carboxylase 总被引:2,自引:0,他引:2
The in vivo induction of rat liver acetyl-CoA carboxylase (ACC) the rate-limiting enzyme of fatty acid biosynthesis, has been examined by immunoblotting, avidin blotting, and enzyme isolation. Three high-molecular-weight immunoreactive bands (Mr 220,000-260,000) were recognized in liver extracts by an anti-carboxylase polyclonal antiserum. Two bands, A and B, comigrated on sodium dodecyl sulfate polyacrylamide gels with purified acetyl-CoA carboxylase, were avidin binding, and were dramatically induced following high carbohydrate refeeding. Only band A was recognized on immunoblots using a monoclonal antibody directed against acetyl-CoA carboxylase, suggesting that band B is a proteolytic fragment in which the epitope recognized by the monoclonal antibody is absent. Following refeeding, approximately 57% of acetyl-CoA carboxylase mass (band A + band B) was present in the high-speed supernatant fraction, while 34 and 9% were in the high-speed (microsomal) and low-speed pellet fractions, respectively. Refeeding caused a large increase in total acetyl-CoA carboxylase mass, the magnitude of which differed in the various fractions. In the low-speed supernatant, a 20-fold increase in ACC mass was observed, while a 12-fold increase was seen in the high-speed supernatant. The fold increase in the high-speed pellet was even greater (greater than 27-fold). Acetyl-CoA carboxylase purified by avidin-Sepharose chromatography from fasted/refed rats had an approximate 4-fold higher Vmax and a significantly lower Ka for citrate than enzyme purified from fasted animals. The results of this study indicate that the induction of hepatic ACC that occurs during high carbohydrate refeeding of the fasted rat predominantly involves increases in enzyme content in both cytosol and microsomes, but is also accompanied by an increase in enzyme specific activity. 相似文献
11.
The effects of citrate and cyclic AMP on the rate and degree of phosphorylation and inactivation of rat liver acetyl-CoA carboxylase were examined. High citrate concentrations (10 to 20 mM), which are generally used to stabilize and activate the enzyme, inhibit phosphorylation and inactivation of carboxylase. At lower concentrations of citrate, the rate and degree of phosphorylation are increased. Furthermore, phosphorylation and enzyme inactivation are affected by cyclic AMP under these conditions. At high citrate concentrations, cyclic AMP has little or no effect on inactivation and phosphorylation of acetyl-CoA carboxylase. Phosphorlation and inactivation of carboxylase is accompanied by depolymerization of the polymeric form of the enzyme into intermediate and protomeric forms. Depolymerization of carboxylase requires the transfer of the gamma-phosphate group from ATP to carboxylase. Inactivation occurs in the absence of CO2, which indicates that phosphorylation of the enzyme is the cause of inactivation and depolymerization, i.e. carboxylation of the enzyme is not responsible for inactivation of the enzyme. 相似文献
12.
Trang Pham Elizabeth Walden Sylvain Huard John Pezacki Morgan D Fullerton Kristin Baetz 《Genetics》2022,221(4)
Acetyl-CoA Carboxylase 1 catalyzes the conversion of acetyl-CoA to malonyl-CoA, the committed step of de novo fatty acid synthesis. As a master regulator of lipid synthesis, acetyl-CoA carboxylase 1 has been proposed to be a therapeutic target for numerous metabolic diseases. We have shown that acetyl-CoA carboxylase 1 activity is reduced in the absence of the lysine acetyltransferase NuA4 in Saccharomyces cerevisiae. This change in acetyl-CoA carboxylase 1 activity is correlated with a change in localization. In wild-type cells, acetyl-CoA carboxylase 1 is localized throughout the cytoplasm in small punctate and rod-like structures. However, in NuA4 mutants, acetyl-CoA carboxylase 1 localization becomes diffuse. To uncover mechanisms regulating acetyl-CoA carboxylase 1 localization, we performed a microscopy screen to identify other deletion mutants that impact acetyl-CoA carboxylase 1 localization and then measured acetyl-CoA carboxylase 1 activity in these mutants through chemical genetics and biochemical assays. Three phenotypes were identified. Mutants with hyper-active acetyl-CoA carboxylase 1 form 1 or 2 rod-like structures centrally within the cytoplasm, mutants with mid-low acetyl-CoA carboxylase 1 activity displayed diffuse acetyl-CoA carboxylase 1, while the mutants with the lowest acetyl-CoA carboxylase 1 activity (hypomorphs) formed thick rod-like acetyl-CoA carboxylase 1 structures at the periphery of the cell. All the acetyl-CoA carboxylase 1 hypomorphic mutants were implicated in sphingolipid metabolism or very long-chain fatty acid elongation and in common, their deletion causes an accumulation of palmitoyl-CoA. Through exogenous lipid treatments, enzyme inhibitors, and genetics, we determined that increasing palmitoyl-CoA levels inhibits acetyl-CoA carboxylase 1 activity and remodels acetyl-CoA carboxylase 1 localization. Together this study suggests yeast cells have developed a dynamic feed-back mechanism in which downstream products of acetyl-CoA carboxylase 1 can fine-tune the rate of fatty acid synthesis. 相似文献
13.
The activities of hepatic acetyl-CoA carboxylase and fatty acid synthetase undergo two distinct types of development in the perinatal chick. The first increase begins prior to hatching, continues after hatching in the starved chick, and is independent of feeding. The second increase is caused by feeding and is reversed by starvation (A. G. Goodridge (1973) J. Biol. Chem.248, 1932–1938). We have purified these enzymes to homogeneity and raised antibodies to them in rabbits. Using immunochemical techniques we have established that the activity changes in both types of development were a function of changes in the concentrations of enzyme proteins. All activity changes were accompanied by similar changes in the relative rates of synthesis of the two enzymes. Regulation of the activities of acetyl-CoA carboxylase and fatty acid synthetase was further characterized in liver cells from 19-day-old embryos maintained in culture in a chemically defined medium. After 3 days in culture in the absence of hormones, the activities of the enzymes increased significantly with respect to the activities of the freshly prepared cells. Addition of either insulin or triiodothyronine alone caused additional small increases. Insulin plus triiodothyronine caused 8- and 15-fold increases in acetyl-CoA carboxylase and fatty acid synthetase, respectively, relative to cells incubated without hormones. In the presence of insulin alone glucagon had no effect on the activity of either enzyme. In the presence of insulin plus triiodothyronine, glucagon inhibited the increase in enzyme activities by about 75%. The results of quantitative immunoprecipitin tests indicated that activity changes caused by the various hormones were functions of changes in the concentrations of the enzyme proteins. The effects of the hormones on enzyme activities were accompanied by comparable or larger changes in the relative rates of synthesis of the enzymes. Under a wide variety of experimental conditions, both in vivo and in culture, the relative rates of synthesis of acetyl-CoA carboxylase and fatty acid synthetase are regulated coordinately. Under some of these conditions, synthesis of malic enzyme also is regulated coordinately with the syntheses of acetyl-CoA carboxylase and fatty acid synthetase. The common intracellular mechanisms underlying the coordinate control remain to be elucidated. 相似文献
14.
Chick liver cell monolayers synthesize fatty acids at in vivo rates and are responsive to insulin and glucagon. High rates of fatty acid synthesis are maintained with insulin present and lost slowly without insulin. Glucagon or 3',5'-cyclic AMP cause immediate cessation of fatty acid synthesis. The site of inhibition appears to be cytoplasmic acetyl-CoA carboxylase which catalyzes the first committed step of fatty acid synthesis. Liver carboxylase exists either as catalytically inactive protomers or active filamentous polymers. Citrate, an allosteric activator of the enzyme, is required for both catalysis and polymerization. Glucagon and cAMP cause an immediate decrease in the cytoplasmic citrate concentration of chick liver cells apparently by inhibiting the conversion of glucose to citrate at the phosphofructokinase reaction. Since fatty acid synthesis and citrate level are closely correlated, citrate appears to be a feed-forward activator of the carboxylase in vivo. Compelling evidence indicates that carboxylase filaments are present in the intact cell when citrate levels are high and depolymerize when citrate levels fall. Hence, carboxylase activity and fatty acid synthetic rate appear to be determined by cytoplasmic citrate level. 相似文献
15.
When fasted rats were refed for 4 days with a carbohydrate and protein diet, a carbohydrate diet (without protein) or a protein diet (without carbohydrate), the effects of dietary nutrients on the fatty acid synthesis from injected tritiated water, the substrate and effector levels of lipogenic enzymes and the enzyme activities were compared in the livers. In the carbohydrate diet group, although acetyl-CoA carboxylase was much induced and citrate was much increased, the activity of acetyl-CoA carboxylase extracted with phosphatase inhibitor and activated with 0.5 mM citrate was low in comparison to the carbohydrate and protein diet group. The physiological activity of acetyl-CoA carboxylase seems to be low. In the protein diet group, the concentrations of glucose 6-phosphate, acetyl-CoA and malonyl-CoA were markedly higher than in the carbohydrate and protein group, whereas the concentrations of oxaloacetate and citrate were lower. The levels of hepatic cAMP and plasma glucagon were high. The activities of acetyl-CoA carboxylase and also fatty acid synthetase were low in the protein group. By feeding fat, the citrate level was not decreased as much as the lipogenic enzyme inductions. Comparing the substrate and effector levels with the Km and Ka values, the activities of acetyl-CoA carboxylase and fatty acid synthetase could be limited by the levels. The fatty acid synthesis from tritiated water corresponded more closely to the acetyl-CoA carboxylase activity (activated 0.5 mM citrate) than to other lipogenic enzyme activities. On the other hand, neither the activities of glucose-6-phosphate dehydrogenase and malic enzyme (even though markedly lowered by diet) nor the levels of their substrates appeared to limit fatty acid synthesis of any of the dietary groups. Thus, it is suggested that under the dietary nutrient manipulation, acetyl-CoA carboxylase activity would be the first candidate of the rate-limiting factor for fatty acid synthesis with the regulations of the enzyme quantity, the substrate and effector levels and the enzyme modification. 相似文献
16.
Zonation of hepatic lipogenic enzymes identified by dual-digitonin-pulse perfusion. 总被引:2,自引:2,他引:0
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The zonal distribution within rat liver of acetyl-CoA carboxylase, ATP citrate-lyase and fatty acid synthase, the principal enzymes of fatty acid synthesis, was investigated by using dual-digitonin-pulse perfusion. Analysis of enzyme mass by immunoblotting revealed that, in normally feeding male rats, the periportal/perivenous ratio of acetyl-CoA carboxylase mass was 1.9. The periportal/perivenous ratio of ATP citrate-lyase mass was 1.4, and fatty acid synthase exhibited the largest periportal/perivenous mass gradient, having a ratio of 3.1. This pattern of enzyme distribution was observed in male rats only; in females, the periportal/perivenous ratio of enzyme mass was nearly equal. The periportal/perivenous gradients for acetyl-CoA carboxylase, ATP citrate-lyase and fatty acid synthase observed in fed (and fasted) males were abolished when animals were fasted (48 h) and refed (30 h) with a high-carbohydrate/low-fat diet. As determined by enzyme assay of eluates obtained from the livers of normally feeding male rats, there is also periportal zonation of acetyl-CoA carboxylase activity, expressed either as units per mg of eluted protein or units per mg of acetyl-CoA carboxylase protein, suggesting the existence of gradients in both enzyme mass and specific activity. From these results, we conclude that the enzymes of fatty acid synthesis are zonated periportally in the liver of the normally feeding male rat. 相似文献
17.
Acute effects in vivo of anti-insulin serum on rates of fatty acid synthesis and activities of acetyl-coenzyme A carboxylase and pyruvate dehydrogenase in liver and epididymal adipose tissue of fed rats. 总被引:5,自引:0,他引:5
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Plasma insulin concentrations in fed rats were altered acutely by administration of glucose or anti-insulin serum. Rates of fatty acid synthesis in adipose tissue and liver were estimated from the incorporation of 3H from 3H2O. In the adipose tissue dehydrogenase and acetyl-CoA carboxylase were evident. In liver, although changes in rates of fatty acid synthesis were found, the initial activity of pyruvate dehydrogenase did not alter, but small parallel changes in acetyl-CoA carboxylase activity were observed. 相似文献
18.
《Biochimica et Biophysica Acta (BBA)/Lipids and Lipid Metabolism》1981,663(3):585-597
In agreement with its well-known inhibition of mitochondrial carrier-mediated pyruvate transport, α-cyano-4-hydroxycinnamate elevates pyruvate and lactate levels in suspensions of isolated rat hepatocytes, whereas it lowers citrate levels and causes strongly depressed rates of fatty acid synthesis with glucose as carbon precursor. It stimulates the oxidation of exogenous fatty acids and inhibits their esterification.α-Cyano-4-hydroxycinnamate also impairs fatty acid synthesis from substrates (acetate, octanoate) that bypass mitochondrial pyruvate transport. Cholesterol synthesis from acetate, a process utilizing the same cytosolic acetyl-CoA pool as does fatty acid synthesis, is hardly affected by α-cyano-4-hydroxy-cinnamate. These observations suggest an inhibitory site of action of α-cyano-4-hydroxycinnamate located in the fatty-acid biosynthetic pathway itself. This suggestion has been confirmed by demonstrating the inhibition of purified rat-liver acetyl-CoA carboxylase by α-cyano-4-hydroxycinnamate at concentrations prevailing in the intact cell upon incubation with this compound.Maximal inhibition of purified acetyl-CoA carboxylase requires about 20 min of preincubation of the enzyme with α-cyano-4-hydroxycinnamate. Fatty acid synthesis from acetate in the intact cells is further diminished after an incubation time of 20 min.The inhibition by α-cyano-4-hydroxycinnamate of fatty acid synthesis from acetate can be partially overcome by insulin. Possible interactions of the inhibitor and the hormone at the level of acetyl-CoA carboxylase are discussed.It is concluded that α-cyano-4-hydroxycinnamate does not provide a simple and unequivocal tool to distinguish between actions of effectors on hepatic fatty acid synthesis per se and on the glycolytic pathway. 相似文献
19.
Troglitazone stimulates acetyl-CoA carboxylase activity through a post-translational mechanism 总被引:1,自引:0,他引:1
Troglitazone, a thiazolidinedione, is known to act as an insulin sensitizer. The various effects of the drug include stimulation of glucose utilization and inhibition of gluconeogenesis and fatty acid oxidation. We studied the effect of troglitazone treatment on rat liver acetyl-CoA carboxylase (ACC), the key enzyme that catalyzes the formation of malonyl-CoA, the rate-limiting step in the synthesis of long chain fatty acids. Treatment of rats with troglitazone for 18 days resulted in more than 200% increase in the activity of hepatic acetyl-CoA carboxylase (1.01+/-0.14 and 2.33+/-0.28 mU/mg supernatant protein for control and troglitazone-treated rats, respectively) (p<0.001). The expression of acetyl-CoA carboxylase mRNA, as studied by RNAse protection assay, was not significantly different between the two groups of animals. The ACC from control and troglitazone-treated groups was purified by avidin-affinity chromatography. The purified enzyme migrated as a major protein band (Mr 262,000) on SDS-polyacrylamide gels. Troglitazone treatment was associated with increased citrate sensitivity of ACC. The specific activity of the purified preparation in troglitazone-treated rats was increased by 67% (2.5 vs. 1.5 U/mg). Quantitation of alkali-labile phosphate content of the purified preparation revealed 5.66+/-0.17 and 6.29+/-0.13 mol Pi/mol subunit of 262 Kda for control and troglitazone-treated rats, respectively (P<0.01). The subtle increase in phosphate content does not explain the observed activation of the enzyme. It is possible that additional mechanisms such as troglitazone related rearrangement of the occupancy of select phosphate binding sites or altered binding of the biotin cofactor may also contribute to the observed activation of ACC. 相似文献
20.
De novo fatty acid synthesis in developing rat lung 总被引:1,自引:0,他引:1
The rate of de novo fatty acid synthesis in developing rat lung was measured by the rate of incorporation of 3H from 3H2O into fatty acids in lung slices and by the activity of acetyl-CoA carboxylase in fetal, neonatal and adult lung. Both tritium incorporation and acetyl-CoA carboxylase activity increased sharply during late gestation, peaked on the last fetal day, and declined by 50% 1 day after birth. In the adult, values were only one-half the peak fetal rates. In vitro regulation of acetyl-CoA carboxylase activity in fetal lung was similar to that described in adult non-pulmonary tissues: activation by citrate and inhibition by palmitoyl-CoA. Similarly, incubation conditions that favored enzyme phosphorylation inhibited acetyl-CoA carboxylase activity in lung while dephosphorylating conditions stimulated activity. Incorporation of [U-14 C]glucose into lung lipids during development was influenced heavily by incorporation into fatty acids, which generally paralleled the rate of tritium incorporation into fatty acids. The relative utilization of acetyl units from exogenous glucose for overall fatty acid synthesis was greater in adult lung than in fetal or neonatal lung, suggesting that other substrates may be important for fatty acid synthesis in developing lung. In fetal lung explants, de novo fatty acid synthesis was inhibited by exogenous palmitate. Taken together, these data suggest that de novo synthesis may be an important source of saturated fatty acids in fetal lung but of lesser importance in the neonatal period. Furthermore, the regulation of acetyl-CoA carboxylase activity and fatty acid synthesis in lung may be similar to non-pulmonary tissues. 相似文献