Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:6.4.1.1 (pyruvate carboxylase)
1,516 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Insulin was found to double the rate of incorporation of H14CO3- into protein by segments of rat epididymal adipose tissue provided the incubation medium contained a suitable energy substrate such as fructose. Overall protein synthesis was increased by insulin to a lesser extent, one-third as measured by tritiated water indicating that insulin also increased CO2 fixation into amino acids. The latter could be demonstrated only when the tissue amino acid pools were expanded by the addition of aspartate to the incubation medium. The pattern of labeling observed in the amino acids indicated that CO2 fixation occurred primarily at the pyruvate carboxylase step. Addition of pyruvate to the incubation medium also increased CO2 fixation and this effect was not additive with that of insulin, suggesting that insulin acted by increasing the availability of pyruvate to the carboxylase. No change in carboxylase activity could be measured. Mitochondria isolated from tissue exposed to insulin retained a higher capacity to fix CO2 into acid-soluble products provided they were not freeze-thawed or sonicated. Uptake of pyruvate by mitochondria incubated 1 min at 2 degrees C or 5 s at 15 degrees C was doubled by prior insulin treatment of the tissue. It is concluded that insulin increases the flux through pyruvate carboxylase in adipose tissue in part by increasing the transport of pyruvate through the inner mitochondrial membrane.
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PMID:Effects of insulin on CO2 fixation in adipose tissue. Evidence for regulation of pyruvate transport. 640 Dec 94

Administration of physiologically low doses of bicarbonate into rats caused an inhibition of oxidative phosphorylation and a transitory decrease in ATP content in liver mitochondria; at the same time, concentrations of malate and glutamate were unaltered and those of pyruvate and phosphoenolpyruvate were decreased. Insulin removed the bicarbonate effect on mitochondrial functions but affected only slightly the distribution of metabolites. Bicarbonate appears to activate pyruvate carboxylase and to inhibit succinate dehydrogenase as well as the operation of tricarboxylic acid cycle due to accumulation of oxaloacetate. The effect of insulin mimics acceleration of decarboxylation reactions in mitochondria.
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PMID:[Effect of bicarbonate and insulin on energy metabolism in the mitochondria of rat liver]. 676 May 41

Previous studies indicated that in pancreatic islets the amount of glucose-derived pyruvate that enters mitochondrial metabolism via carboxylation is approximately equal to that entering via decarboxylation and that both carboxylation and decarboxylation are correlated with capacitation of glucose metabolism and insulin release. The relatively high rate of carboxylation is consistent with the current study's finding that pyruvate carboxylase is as abundant in pancreatic islets as it is in liver and kidney. Since islets do not contain phosphoenolpyruvate carboxykinase and, therefore, cannot carry out glyconeogenesis from pyruvate, the carboxylase might be present in the islet to participate in novel anaplerotic reactions. This idea was first explored by incubating mitochondria from various tissues with pyruvate. Mitochondria from tissues, such as pancreatic islets, liver, and kidney, in which pyruvate carboxylase is abundant, exported a large amount of malate and little or no citrate, isocitrate, and aspartate to the medium. The amount of malate within the mitochondria was < 1% that in the medium. When pancreatic islet mitochondria were incubated with [1-14C]pyruvate, radioactive carbon appeared in the medium primarily in malate. Very little radioactivity appeared in amino acids, and little or no radioactivity appeared in citrate and isocitrate. Carbon 1 of pyruvate can be incorporated into malate and other citric acid cycle intermediates only via carboxylation, as this carbon would be lost via decarboxylation when pyruvate enters the citric acid cycle as acetyl-CoA via the pyruvate dehydrogenase reaction. The amount of malate formed equaled the 14CO2 formed and the radioactivity from C-1 of pyruvate recovered in malate slightly exceeded the formation of 14CO2 in agreement with our previous studies that reported a high rate of carboxylation of pyruvate in intact islets. When intact pancreatic islets were incubated with methyl [U-14C]succinate as a mitochondrial source of four-carbon dicarboxylic acids, radioactivity appeared in pyruvate and lactate. Taken together with previous studies, the current results suggest that during glucose-induced insulin secretion there is a shuttle operating across the mitochondrial membrane in which glucose-derived pyruvate is taken up by mitochondria and carboxylated to oxaloacetate by pyruvate carboxylase. The oxaloacetate is converted to malate which exits the mitochondrion, where, in the cytosol, it is decarboxylated to pyruvate in the reaction catalyzed by malic enzyme. This pyruvate re-enters mitochondrial pools. Such a cycle produces NADPH in the cytosol. Since it is a cycle, this shuttle can produce far more NADPH than the pentose phosphate pathway, which is known to be a very minor route of glucose metabolism in the islet.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Feasibility of a mitochondrial pyruvate malate shuttle in pancreatic islets. Further implication of cytosolic NADPH in insulin secretion. 765 22

Pancreatic islets were cultured for 1 day in the presence of 1 to 20 mM glucose and islet proteins were separated on polyacrylamide gels and transferred to nitrocellulose. Pyruvate carboxylase and an unidentified biotin-containing protein were visualized with [125I]streptavidin followed by autoradiography. The amount of pyruvate carboxylase was proportional to the concentration of glucose. Estimates of the amount of the enzyme in islets were made by comparing the density of the islet pyruvate carboxylase band with a standard curve of various amounts of authentic pyruvate carboxylase. This indicated that the enzyme comprised 0.4% of total islet protein. Net synthesis of the enzyme was increased by cAMP and methyl succinate. A nuclear run-on assay showed that glucose caused increases in pyruvate carboxylase and pyruvate dehydrogenase E1 alpha subunit transcripts and decreases in branched chain ketoacid dehydrogenase E1 alpha transcripts in rat insulinoma (RINm5F) cells. Pancreatic islets cultured in the presence of 1 mM glucose for 1 day cannot respond to glucose with insulin release. Previous studies demonstrated that carbon flux into the citric acid cycle intermediates via both carboxylation and decarboxylation is decreased in glucose-incapacitated islets (M. J. MacDonald, 1993, Arch. Biochem. Biophys. 300, 205-214), 1993). The current results support the idea that carboxylation of glucose-derived pyruvate, as well as decarboxylation of pyruvate, is important for glucose-induced insulin secretion.
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PMID:Influence of glucose on pyruvate carboxylase expression in pancreatic islets. 777 76

When 3T3-L1 fibroblasts differentiate to adipocytes, the specific activity of pyruvate carboxylase (PC) increases about 25-fold in parallel with its intracellular protein concentration. The increase in PC protein concentration is accompanied by a 9-10-fold increase in the relative abundance of 4.2 kb PC mRNA measured by Northern-blot analysis using a cDNA probe encoding a segment of the PC gene of 3T3-L1 adipocytes. The effects of cyclic AMP (cAMP) alone and together with insulin on levels of cellular protein, PC activity, PC protein and on the relative abundance of PC mRNA were examined in mature 3T3-L1 adipocytes. Adipocytes exposed to cAMP for 24 h exhibited a 25% decrease in cellular protein and marked decreases in enzyme activity (88%) and PC mRNA abundance (98%) compared with untreated adipocyte controls. After 48 h of exposure to cAMP, PC activity and PC mRNA diminished to levels approaching their detection limits. When exposed to medium containing cAMP plus insulin, adipocyte enzyme activity and PC mRNA declined more slowly during the first 24 h exposure (about 20% decrease) but after 48 h fell to values comparable with those of adipocytes exposed to cAMP alone. Despite these decreases in enzyme activity, the PC protein content of adipocytes treated with cAMP alone or cAMP plus insulin are nearly identical with that of control adipocytes. The inactivation of PC in cAMP-treated adipocytes does not involve loss of the prosthetic group from the holoenzyme. Cross-linking experiments suggest that the spatial arrangement of protomers in inactive PC may differ from that in the active tetrameric enzyme. Data presented suggest that, in addition to inducing inactivation, cAMP may also regulate adipocyte PC by decreasing transcription of the PC gene and/or enhancing the rate of degradation of PC mRNA.
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PMID:Regulation of pyruvate carboxylase in 3T3-L1 cells. 786 11

In humans, endurance training markedly reduces the rate of hepatic glucose production during exercise. To determine whether this is due to a reduction in glycogenolysis, in gluconeogenesis, or in both processes, six men were studied at rest and during 2 h of cycle ergometer exercise at 60% pretraining peak O2 consumption (VO2peak), both before and after completion of a strenuous endurance training program (cycling at 75-100% VO2peak for 45-90 min/day, 6 days/wk for 12 wk). The overall rate of glucose appearance (Ra) was determined using a primed continuous infusion of [6,6-2H]glucose, whereas the rate of gluconeogenesis (Rgng) was estimated from the incorporation of 13C into glucose (via pyruvate carboxylase) from simultaneously infused [13C]bicarbonate. Training did not affect glucose kinetics at rest but reduced the average Ra during exercise by 42% [from 36.8 +/- 3.8 to 21.5 +/- 3.6 (SE) mumol.min-1.kg-1; P < 0.001]. This decrease appeared to be mostly due to a reduction in hepatic glycogenolysis. However, the estimated Rgng during exercise also decreased significantly (P < 0.001) with training, falling from 7.5 +/- 1.6 mumol.min-1.kg-1 (23 +/- 3% of total Ra) before training to 3.1 +/- 0.6 mumol.min-1.kg-1 (14 +/- 3% of total Ra) after training. These training-induced adaptations in hepatic glucose metabolism were associated with an attenuated hormonal response to exercise (i.e., higher insulin and lower glucagon, norepinephrine, and epinephrine concentrations) as well as a reduced availability of gluconeogenic precursors (i.e., lower lactate and glycerol concentrations). We conclude that endurance training reduces both hepatic glycogenolysis and gluconeogenesis during prolonged exercise in men.
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PMID:Effect of endurance training on hepatic glycogenolysis and gluconeogenesis during prolonged exercise in men. 790 Jul 83

Hepatocyte monolayers from neonatal calves were used to determine the effects of glucagon and insulin on incorporation of carbon from [2-14C]propionate, [1-14C]lactate, [U-14C]lactate, and [1,3-14C]glycerol into glucose and glycogen. Glucagon increased gluconeogenesis (nmol substrate incorporated into glucose or glycogen.micrograms DNA-1.h-1) from propionate and lactate but not from glycerol. Insulin decreased gluconeogenesis from [2-14C]propionate but was without effect on gluconeogenesis from [U-14C]lactate or [1,3-14C]glycerol. Net de novo glycogenesis (nmol substrate retained in cell glycogen.micrograms DNA-1.h-1) from propionate, lactate, and glycerol was decreased by glucagon and increased by insulin. Glucagon effects on gluconeogenesis, but not glycogenesis, were mimicked by dibutyryl adenosine 3',5'-cyclic monophosphate. Lactate flux through pyruvate carboxylase accounts for > or = 91% of lactate carbon flux to glucose, and this proportion was unchanged by glucagon or insulin. Gluconeogenesis from propionate and lactate is regulated by substrate concentration and glucagon in bovine hepatocyte monolayers. The data indicate that, in neonatal bovine liver, glucagon acts on a process common to lactate and propionate to increase gluconeogenesis, and insulin opposes these effects on gluconeogenesis from propionate but not lactate.
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PMID:Regulation of gluconeogenesis by insulin and glucagon in the neonatal bovine. 818 66

Carbonic anhydrase (CA) was examined in two adipocyte cell lines, 3T3-L1 and 3T3-F442A. Both CA III and non-CA III activities, measured by 18O mass spectrometry, were present in 3T3-L1 and 3T3-F442A adipocytes; however, no CA activity was detected in 3T3 preadipocytes of either line. These observations were supported by immunoblot experiments employing CA III and CA II isoform-specific antisera. CA III, a major protein in rodent and murine adipocytes, and CA II, another isoform known to be present in adipose tissue, were observed only in the differentiated 3T3 adipocytes. The differentiation-dependent expression of these isozymes may imply an adipocyte-related role for CA. Compared with cultures maintained in the absence of insulin, 3T3 adipocytes maintained in the presence of insulin exhibited 65-90% lower concentrations of CA III. CA II was unaffected. This negative effect of insulin on CA III may explain the metabolic regulation of adipose CA III observed in vivo. After media changes, 3T3 adipocyte cultures rapidly lower media pH, which in turn lowers the bicarbonate/CO2 of bicarbonate/CO2-buffered media. Cultures maintained at low pH displayed 50-90% lower concentrations of CA II and CA III. Similarly, cultures maintained in a low bicarbonate/CO2 media (GibCO2-I medium containing 1 mM bicarbonate under an atmosphere of 100% humidified air) displayed 30-50% lower CA II and CA III concentrations. Thus CA II and CA III concentrations are influenced by pH and bicarbonate/CO2. Neither effect, the pH or the GibCO2-I media effect, was associated with changes in the concentration of pyruvate carboxylase or ATP citrate lyase (2 markers of adipocyte differentiation). Because the regulation by pH and bicarbonate/CO2 may be relatively selective for CA in adipocytes, a simple method for reducing the concentration/activity of CA in 3T3 adipocytes is described that may be a useful tool for studies on the physiological role of the enzyme.
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PMID:Differentiation-dependent expression of carbonic anhydrase II and III in 3T3 adipocytes. 833 33

Pancreatic islets were cultured for 24 h in the presence of 1 mM glucose, which renders islets incapable of responding to glucose with insulin release. These islets were compared to islets maintained at 20 mM glucose for 24 h. Detritiation of [2-3H]glucose and [5-3H]glucose in 1 mM glucose islets was normal, suggesting that glucose transport and phosphorylation and all enzymes of glycolysis were not down-regulated in the incapacitated islets. 14CO2 formation from [U-14C]glucose and [6-14C]glucose was inhibited up to 80% and 14CO2 from methyl succinate was inhibited up to 60%, indicating that down-regulation at (a) mitochondrial site(s) might explain the incapacitated insulin release. 14CO2 formation from [3,4-14C]glucose (which becomes [1-14C]pyruvate) was decreased, indicating that the reaction catalyzed by pyruvate dehydrogenase was down-regulated. This decrease, however, was not as large as the decreases in 14CO2 formation from [U-14C]glucose, [2-14C]glucose (which becomes [2-14C]pyruvate), or [6-14C]glucose (which becomes [3-14C]pyruvate), indicating that other reactions were also down-regulated. 14CO2 formation from [1-14C]glucose was inhibited less than that from [6-14C]glucose in the incapacitated islets (34 vs 54%) and these rates indicated that flux of glucose through the pentose phosphate pathway was increased in the incapacitated islet, such that 29% (0.4 nmol of 1.4 glucose/100 islets/90 min) was metabolized via this pathway in the incapacitated islet but only 3.4% (0.1 of 2.9 nmol glucose/100 islets/90 min) was metabolized via the pentose pathway in the 20 mM glucose islets. With rates of 14CO2 evolved from glucose labeled at C2 and C6 and from methyl succinate labeled at C1 + C4 and C2 + C3 the 14CO2 ratio formula was used to calculate the ratios of carboxylated and decarboxylated pyruvate. Roughly equal amounts of pyruvate entered the citric acid cycle by each route in islets maintained for 24 h at 1, 5, or 20 mM glucose. The results indicate that decarboxylation and carboxylation of pyruvate were about equally suppressed in incapacitated islets and that direct inhibition of reactions of the cycle was unlikely. This is consistent with evidence which indicates that down-regulation of both pyruvate carboxylase and pyruvate dehydrogenase occurs in incapacitated islets, i.e., under long-term conditions that modify amounts of enzymes (MacDonald et al., 1991, J. Biol. Chem. 266, 22392-22397).(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Estimates of glycolysis, pyruvate (de)carboxylation, pentose phosphate pathway, and methyl succinate metabolism in incapacitated pancreatic islets. 837 57

Previous work demonstrated that methyl esters of succinate are potent insulin secretagogues in pancreatic islets, while unesterified succinate is not. This can be explained by studies reported here, which show that 14C-labeled dimethyl succinate is metabolized to 14CO2 by pancreatic islets, but that 14C-labeled succinic acid is not metabolized. Islets maintained at 1 mM glucose in tissue culture medium for 1 day lose the ability to release insulin in response to glucose and glucose metabolism is decreased 50-80%. The metabolism of dimethyl [1,4-14C]succinate and dimethyl [2,3-14C]succinate is decreased 50-60% in these incapacitated islets relative to islets maintained at 20 mM glucose. From the ratio of 14CO2 formed from dimethyl [1,4-14C]succinate, relative to that from dimethyl [2,3-14C]succinate, "acetate" ratios of 4.9-6.2 were calculated and from the ratio of 14CO2 formed from [2-14C]glucose, relative to that from [6-14C]glucose, "pyruvate ratios" of 1.6-1.7 were calculated. According to the 14CO2 ratios method, these ratios indicate that 53-66% of pyruvate derived from glucose enters the citric acid cycle via carboxylation and 34-47% enters via decarboxylation. Malic enzyme, which carboxylates pyruvate in the cytosol, was normal in islets maintained at 1 mM glucose. Previous work indicated that inhibition of glucose metabolism in islets maintained at low glucose is due to decreased net synthesis of the mitochondrial enzymes pyruvate dehydrogenase and pyruvate carboxylase [J. Biol. Chem. (1991) 266, 22392-22397], which decarboxylate and carboxylate pyruvate, respectively. Acetate (1 mM) but not pyruvate, when added to islets maintained at low glucose, increased dimethyl succinate metabolism to almost that of islets maintained at high glucose. This is consistent with a low amount of pyruvate dehydrogenase being unable to supply acetyl-CoA for condensation with oxalacetate (derived from succinate) and that the rate of the citric acid cycle could be enhanced by adding acetate which can bypass the reaction catalyzed by pyruvate dehydrogenase.
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PMID:Metabolism of the insulin secretagogue methyl succinate by pancreatic islets. 842 53


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