Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Pyruvates”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 343 records · Page 19Linked to original sources
PubMed1976

The elementary reactions of the pig heart pyruvate dehydrogenase complex. A study of the inhibition by phosphorylation.

1. A method was devised for preparing pig heart pyruvate dehydrogenase free of thiamin pyrophosphate (TPP), permitting studies of the binding of [35S]TPP to pyruvate dehydrogenase and pyruvate dehydrogenase phosphate. The Kd of TPP for pyruvate dehydrogenase was in the range 6.2-8.2 muM, whereas that for pyruvate dehydrogenase phosphate was approximately 15 muM; both forms of the complex contained about the same total number of binding sites (500 pmol/unit of enzyme). EDTA completely inhibited binding of TPP; sodium pyrophosphate, adenylyl imidodiphosphate and GTP, which are inhibitors (competitive with TPP) of the overall pyruvate dehydrogenase reaction, did not appreciably affect TPP binding. 2. Initial-velocity patterns of the overall pyruvate dehydrogenase reaction obtained with varying TPP, CoA and NAD+ concentrations at a fixed pyruvate concentration were consistent with a sequential three-site Ping Pong mechanism; in the presence of oxaloacetate and citrate synthase to remove acetyl-CoA (an inhibitor of the overall reaction) the values of Km for NAD+ and CoA were 53+/- 5 muM and 1.9+/-0.2 muM respectively. Initial-velocity patterns observed with varying TPP concentrations at various fixed concentrations of pyruvate were indicative of either a compulsory order of addition of substrates to form a ternary complex (pyruvate-Enz-TPP) or a random-sequence mechanism in which interconversion of ternary intermediates is rate-limiting; values of Km for pyruvate and TPP were 25+/-4 muM and 50+/-10 nM respectively. The Kia-TPP (the dissociation constant for Enz-TPP complex calculated from kinetic plots) was close to the value of Kd-TPP (determined by direct binding studies). 3. Inhibition of the overall pyruvate dehydrogenase reaction by pyrophosphate was mixed non-competitive versus pyruvate and competitive versus TPP; however, pyrophosphate did not alter the calculated value for Kia-TPP, consistent with the lack of effect of pyrophosphate on the Kd for TPP. 4. Pyruvate dehydrogenase catalysed a TPP-dependent production of 14CO2 from [1-14C]pyruvate in the absence of NAD+ and CoA at approximately 0.35% of the overall reaction rate; this was substantially inhibited by phosphorylation of the enzyme both in the presence and absence of acetaldehyde (which stimulates the rate of 14CO2 production two- or three-fold). 5. Pyruvate dehydrogenase catalysed a partial back-reaction in the presence of TPP, acetyl-CoA and NADH. The Km for TPP was 4.1+/-0.5 muM. The partial back-reaction was stimulated by acetaldehyde, inhibited by pyrophosphate and abolished by phosphorylation. 6. Formation of enzyme-bound [14C]acetylhydrolipoate from [3-14C]pyruvate but not from [1-14C]acetyl-CoA was inhibited by phosphorylation. Phosphorylation also substantially inhibited the transfer of [14C]acetyl groups from enzyme-bound [14C]acetylhydrolipoate to TPP in the presence of NADH. 7...

Acetaldehyde↗
PubMed1997

Substrate utilization and maturation of cumulus cell-enclosed mouse oocytes: evidence that pyruvate oxidation does not mediate meiotic induction.

This study was performed to address the possible role of pyruvate in meiotic induction in mouse oocytes. Cumulus cell-enclosed oocytes from primed, immature mice were cultured in 7.5 microliters microdrops under oil for 9 or 18 h in medium containing 4 mmol hypoxanthine l-1 plus 0.23 mmol pyruvate l-1, 1 mmol pyruvate l-1, or 1 mmol pyruvate l-1 plus 5.5 mmol glucose l-1. When compared with cultures containing 0.23 mmol pyruvate l-1, 1 mmol pyruvate l-1 induced germinal vesicle breakdown, and this was preceded by an increase in pyruvate utilization. Addition of glucose prevented both the increase in pyruvate consumption and the meiotic induction. When different combinations of pyruvate and glucose were tested on oocyte maturation in microdrop cultures, a high concentration of pyruvate or glucose alone was stimulatory to maturation. Addition of the complementary energy substrate prevented the induction of germinal vesicle breakdown and reduced the amount of substrate consumption. During spontaneous maturation in vitro, oocyte-cumulus cell complexes consumed glucose for the first 3 h; however, during the second 3 h period, which followed germinal vesicle breakdown, glucose consumption decreased and net pyruvate utilization was initiated. Treatment of hypoxanthine-arrested oocytes with dichloroacetate, an activator of pyruvate dehydrogenase, stimulated pyruvate consumption but had no effect on germinal vesicle breakdown. Although FSH stimulates meiotic resumption, no changes in pyruvate consumption were observed in response to this gonadotrophin. Measurement of oxygen consumption by hypoxanthine-treated complexes revealed no effect of high concentrations of pyruvate on respiration, and FSH treatment resulted in a suppression of oxygen utilization. These data indicate that, in mouse oocyte-cumulus cell complexes, pyruvate and glucose can each modulate metabolism of the other substrate, and the can significantly influence meiotic maturation of the oocyte. In addition, augmentation of pyruvate oxidation does not appear to play a mediating role in meiotic induction triggered by energy substrate manipulation or gonadotrophin treatment.

Animals↗
PubMed1995

Feasibility of a mitochondrial pyruvate malate shuttle in pancreatic islets. Further implication of cytosolic NADPH in insulin secretion.

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)

Animals↗
PubMed1976

Regulation of pyruvate dehydrogenase in rat heart. Mechanism of regulation of proportions of dephosphorylated and phosphorylated enzyme by oxidation of fatty acids and ketone bodies and of effects of diabetes: role of coenzyme A, acetyl-coenzyme A and reduced and oxidized nicotinamide-adenine dinucleotide.

The proportion of active (dephosphorylated) pyruvate dehydrogenase in perfused rat heart was decreased by alloxan-diabetes or by perfusion with media containing acetate, n-octanoate or palmitate. The total activity of the dehydrogenase was unchanged. 2. Pyruvate (5 or 25mM) or dichloroacetate (1mM) increased the proportion of active (dephosphorylated) pyruvate dehydrogenase in perfused rat heart, presumably by inhibiting the pyruvate dehydrogenase kinase reaction. Alloxan-diabetes markedly decreased the proportion of active dehydrogenase in hearts perfused with pyruvate or dichloroacetate. 3. The total activity of pyruvate dehydrogenase in mitochondria prepared from rat heart was unchanged by diabetes. Incubation of mitochondria with 2-oxo-glutarate plus malate increased ATP and NADH concentrations and decreased the proportion of active pyruvate dehydrogenase. The decrease in active dehydrogenase was somewhat greater in mitochondria prepared from hearts of diabetic rats than in those from hearts of non-diabetic rats. Pyruvate (0.1-10 mM) or dichloroacetate (4-50 muM) increased the proportion of active dehydrogenase in isolated mitochondria presumably by inhibition of the pyruvate dehydrogenase kinase reaction. They were much less effective in mitochondria from the hearts of diabetic rats than in those of non-diabetic rats. 4. The matrix water space was increased in preparations of mitochondria from hearts of diabetic rats. Dichloroacetate was concentrated in the matrix water of mitochondria of non-diabetic rats (approx. 16-fold at 10 muM); mitochondria from hearts of diabetic rats concentrated dichloroacetate less effectively. 5. The pyruvate dehydrogenase phosphate phosphatase activity of rat hearts and of rat heart mitochondria (approx. 1-2 munit/unit of pyruvate dehydrogenase) was not affected by diabetes. 6. The rate of oxidation of [1-14C]pyruvate by rat heart mitochondria (6.85 nmol/min per mg of protein with 50 muM-pyruvate) was approx. 46% of the Vmax. value of extracted pyruvate dehydrogenase (active form). Palmitoyl-L-carnitine, which increased the ratio of [acetyl-CoA]/[CoA] 16-fold, inhibited oxidation of pyruvate by about 90% without changing the proportion of active pyruvate dehydrogenase.

Acetates↗
PubMed1977

Studies on inactivation of pyruvate dehydrogenase by palmitoylcarnitine oxidation in isolated rat heart mitochondria.

The oxidation of an optimal concentration of palmitoyl-carnitine, buffered with bovine serum albumin, by isolated rat heart mitochondria was found to give rise to an inactivation of pyruvate dehydrogenase, provided that the concentration of pyruvate present in the mitochondrial incubation was less than 250 muM. The greatest degree of inactivation was found at the lowest pyruvate concentration used, 50 muM, and this concentration was adopted for further studies in which the rate of mitochondrial respiration was varied. This was done by varying the activity of added hexokinase, in the presence of ATP, MgCl2, and glucose, and thus the availability of ADP to the mitochondrion. The pyruvate concentration in the incubation was approximately stabilized by adding pyruvate on the basis of oxygen consumption, with the ratio of pyruvate consumed:O2 consumed determined by trial and error. This device allowed the maintenance of essentially steady pyruvate concentrations and ATP/ADP ratios for at least 5 min, and allowed the pyruvate dehydrogenase interconversion time to approach a steady state. Activities of pyruvate dehydrogenase after 5 or 6 min of respiration were as follows, with values given in nanomoles/min/mg of protein for incubations containing pyruvate as sole substrate, and values for incubations containing pyruvate plus palmitoylcarnitine given in parentheses: State 4, 27 (9); 55% of State 3, 54 (14); 85% of State 3, 73 (28); State 3, 90 (93). Respiratory states are defined by Chance and Williams (1955) J. Biol. Chem. 217, 409-427). Values at earlier time points are also presented so that some idea may be formed of the time course of pyruvate dehydrogenase inactivation. CoASH/acetyl-CoA, NAD+/NADH, and ATP/ADP ratios were measured at the same time points in precisely scaled up incubations. The presence of palmitoylcarnitine in State 4 was found to give essentially no change in NAD+/NADH and ATP/ADP ratios and thus the inactivation of pyruvate dehydrogenase in that state may be attributed to a decreased CoASH/acetyl-CoA ratio. At a respiratory rate of 85% of State 3, palmitoylcarnitine did not change the ATP/ADP ratio, but lowered both CoASH/acetyl-CoA and NAD+/NADH ratios, both of which may contribute to pyruvate dehydrogenase inactivation. In State 3 there was no pyruvate dehydrogenase inactivation, despite a lowered CoASH/acetyl-CoA ratio in the presence of palmitoylcarnitine. It is concluded that ATP/ADP ratio has a pronounced effect on the interconversion of active and inactive pyruvate dehydrogenase, in according with previous work. Moreover, at a given ATP/ADP ratio, the effects of palmitoylcarnitine oxidation on enzyme interconversion are consistent with a mechanism involving the modulation of the interconversion by NAD+/NADH and CoASH/acetyl-CoA ratios...

Animals↗
PubMed1988

Transport of pyruvate by luminal membrane vesicles from pars convoluta and pars recta of rabbit proximal tubule.

The characteristics of renal transport of pyruvate by luminal membrane vesicles from pars convoluta and pars recta of rabbit proximal tubule were studied. It was found that the uptake of pyruvate in these vesicle preparations occurred by means of multiple transport systems. An electrogenic and Na+-requiring system confined to pars convoluta, exists for transport of pyruvate with an intermediate affinity, KA = 0.71 +/- 0.08 mM. In vesicles from pars recta, the uptake of pyruvate was mediated by a dual transport system with a high (KA1 = 0.30 +/- 0.05 mM) and low affinity (KA2 = 5.75 +/- 0.82 mM). The relation of these three pyruvate transport systems to the transport of other monocarboxylates and dicarboxylates was determined by examination of the inhibitory effect of L-lactate, beta-hydroxybutyrate and L-malate on the renal uptake of pyruvate. It was found that L-malate efficiently reduced the uptake of pyruvate by vesicles from pars convoluta, while addition of monocarboxylates (L-lactate, beta-hydroxybutyrate) has no effect on the transport of pyruvate in this region of proximal tubule. Furthermore, it was observed that Na+-dependent uptake of L-malate was specifically inhibited to about the same extent by 1 mM pyruvate. These findings strongly suggest that pyruvate and L-malate are taken up by the same transport system in vesicles from pars convoluta, which is different from that of L-lactate and beta-hydroxybutyrate. Experiments designed to investigate the substrate specificity for the high and low affinity system for pyruvate in pars recta, revealed that monocarboxylates (L-lactate, beta-hydroxybutyrate) substantially inhibit pyruvate transport at the low, but not the high substrate concentration, indicating that all monocarboxylates compete for the same high-affinity transport system. By contrast, L-malate at low concentrations preferentially inhibited the low-affinity system for pyruvate. This observation led us to suggest that L-malate and pyruvate share a common transport system in pars recta with low affinity for pyruvate and high affinity for L-malate.

Animals↗
PubMed1986

Rate-limiting steps for hepatic gluconeogenesis. Mechanism of oxamate inhibition of mitochondrial pyruvate metabolism.

Oxamate, structural analog of pyruvate, inhibits gluconeogenesis from pyruvate or substrates yielding pyruvate. The inhibitory effect is the result of a decreased mitochondrial pyruvate utilization. Although the inhibition of gluconeogenesis is competitive for pyruvate, in isolated mitochondria oxamate displays a mixed type kinetics inhibitory pattern of pyruvate utilization. Evidence is presented indicating that this mixed type pattern of inhibition is the result of the action of oxamate on two different sites: noncompetitive inhibition of pyruvate carboxylation, and competitive inhibition of pyruvate entry into the mitochondria. At concentrations of pyruvate above 0.4 mM, although pyruvate carboxylation is decreased by 40% by oxamate, no detectable effects on the gluconeogenic flux were observed. This finding strongly indicates that pyruvate carboxylase is not an important rate-limiting step for hepatic gluconeogenesis. Thus, the inhibition of gluconeogenesis at low pyruvate concentrations (less than 0.4 mM) seems to be the result of an interaction of oxamate with the mitochondrial pyruvate translocator, indicating that pyruvate transport across the mitochondrial membrane is the first nonequilibrium step in the gluconeogenic pathway when low physiological concentrations of this substrate are utilized.

Alanine↗
PubMed1984

Stimulation of flux through hepatic pyruvate dehydrogenase by 3-mercaptopicolinate.

Isolated hepatocytes from 24-h-starved rats were used to assess the possible effect of the hypoglycaemic agent 3-mercaptopicolinate on flux through the hepatic pyruvate dehydrogenase complex. Increasing the extracellular pyruvate concentration from 1 mM to 2 mM or 5 mM resulted in an increase in flux through pyruvate dehydrogenase and the tricarboxylic acid cycle as measured by 14CO2 evolution from [1-14C]pyruvate and [3-14C]pyruvate. Gluconeogenesis was inhibited by 3-mercaptopicolinate from both 1 mM and 2 mM pyruvate, but significant increases in malate and citrate concentrations only occurred in cells incubated with 1 mM pyruvate. Flux through pyruvate dehydrogenase was stimulated by 3-mercaptopicolinate with 1 mM pyruvate but was unaltered with 2 mM pyruvate. Dichloroacetate stimulated flux through pyruvate dehydrogenase with no effect on gluconeogenesis in the presence of 1 mM pyruvate. There was no effect of 3-mercaptopicolinate, administered in vivo, to 24-h-starved rats on the activity of pyruvate dehydrogenase in freeze-clamped heart or liver tissue, although the drug did decrease blood glucose concentration and increase the blood concentrations of lactate and alanine. Dichloroacetate, administered in vivo to 24-h-starved rats, increased the activity of pyruvate dehydrogenase in freeze-clamped heart and liver, and caused decreases in the blood concentrations of glucose, lactate, and alanine. The results suggest that 3-mercaptopicolinate increases flux through hepatocyte pyruvate dehydrogenase by an indirect mechanism.

Animals↗
PubMed1972

Mechanisms regulating adipose-tissue pyruvate dehydrogenase.

1. Isolated rat epididymal fat-cell mitochondria showed an inverse relationship between ATP content and pyruvate dehydrogenase activity consistent with competitive inhibition of pyruvate dehydrogenase kinase by ADP. At constant ATP concentration pyruvate rapidly activated pyruvate dehydrogenase in fat-cell mitochondria, an observation consistent with inhibition of fat-cell pyruvate dehydrogenase kinase by pyruvate. Pyruvate dehydrogenase in fat-cell mitochondria was also activated by nicotinate (100mum) and by extramitochondrial Na(+) (replacing K(+)) but not by ouabain or insulin. 2. In rat epididymal fat-pads incubated in vitro pyruvate dehydrogenase was activated by addition of insulin in the absence of substrate or in the presence of glucose (10mm) or fructose (10mm). Glucose and fructose activated the dehydrogenase in the absence or in the presence of insulin, and pyruvate also activated in the absence of insulin. It is concluded that extracellular glucose, fructose and pyruvate may activate the dehydrogenase by raising intracellular pyruvate and that insulin may activate the dehydrogenase by some other mechanism. 3. Ouabain (300mum) and medium in which K(+) was replaced by Na(+), activated pyruvate dehydrogenase in epididymal fat-pads. Prostaglandin E(1) (1mug/ml), 5-methylpyrazole-3-carboxylate (10mum) and nicotinate (10mum), which are as effective as insulin as inhibitors of lipolysis and which like insulin lower tissue concentration of cyclic AMP (adenosine 3':5'-cyclic monophosphate), did not activate pyruvate dehydrogenase. Higher concentrations of prostaglandin E(1) (10mug/ml) and nicotinate (100mum) produced some activation of the dehydrogenase. 4. It is concluded that the activation of pyruvate dehydrogenase by insulin is not due to the antilipolytic effect of the hormone and that the action of insulin in lowering adipose-cell concentrations of cyclic AMP does not afford an obvious explanation for the effect of the hormone on pyruvate dehydrogenase. The possibility that the effects of insulin, ouabain and K(+)-free medium may be mediated by Ca(2+) is discussed.

Adenosine Diphosphate↗
PubMed1975

The mitochondrial pyruvate carrier. Kinetics and specificity for substrates and inhibitors.

1. Studies on the kinetics of pyruvate transport into mitochondria by an 'inhibitor-stop' technique were hampered by the decarboxylation of pyruvate by mitochondria even in the presence of rotenone. Decarboxylation was minimal at 6 degrees C. At this temperature the Km for pyruvate was 0.15 mM and Vmax. was 0.54nmol/min per mg of protein; alpha-cyano-4-hydroxycinnamate was found to be a non-competitive inhibitor, Ki 6.3 muM, and phenyl-pyruvate a competitive inhibitor, Ki 1.8 mM. 2. At 100 muM concentration, alpha-cyano-4-hydroxycinnamate rapidly and almost totally inhibited O2 uptake by rat heart mitochondria oxidizing pyruvate. Inhibition could be detected at concentrations of inhibitor as low as 1 muM although inhibition took time to develop at this concentration. Inhibition could be reversed by diluting out the inhibitor. 3. Various analogues of alpha-cyano-4-hydroxycinnamate were tested on rat liver and heart mitochondria. The important structural features appeared to be the alpha-cyanopropenoate group and the hydrophobic aromatic side chain. Alpha-Cyanocinnamate, alpha-cyano-5-phenyl-2,4-pentadienoate and compound UK 5099 [alpha-cyano-beta-(2-phenylindol-3-yl)acrylate] were all more powerful inhibitors than alpha-cyano-4-hydroxycinnamate showing 50% inhibition of pyruvate-dependent O2 consumption by rat heart mitochondria at concentrations of 200, 200 and 50 nM respectively. 4. The specificity of the carrier for its substrate was studied by both influx and efflux experiments. Oxamate, 2-oxobutyrate, phenylpyruvate, 2-oxo-4-methyl-pentanoate, chloroacetate, dichloroacetate, difluoroacetate, 2-chloropropionate, 3-chloropropionate and 2,2-dichloropropionate all exchanged with pyruvate, whereas acetate, lactate and trichloroacetate did not. 5. Pyruvate entry into the mitochondria was shown to be accompanied by the transport of a proton (or by exchange with an OH-ion). This proton flux was inhibited by alpha-cyano-4-hydroxycinnamate and allowed measurements of pyruvate transport at higher temperatures to be made. The activation energy of mitochondrial pyruvate transport was found to be 113 kJ (27 kcal)/mol and by extrapolation the rate of transport of pyruvate at 37 degrees C to be 42 nmol/min per mg of protein. The possibility that pyruvate transport into mitochondria may be rate limiting and involved in the regulation of gluconegenesis is discussed. 6. The transport of various monocarboxylic acids into mitochondria was studied by monitoring proton influx. The transport of dichloroacetate, difluoroacetate and oxamate appeared to be largely dependent on the pyruvate carrier and could be inhibited by pyruvate-transport inhibitors. However, many other halogenated and 2-oxo acids which could exchange with pyruvate on the carrier entered freely even in the presence of inhibitor.

Animals↗
PubMed1976

Studies on the influence of fatty acids on pyruvate dehydrogenase interconversion in rat-liver mitochondria.

1. The effect of fatty acids on the interconversion of pyruvate dehydrogenase between its active (nonphosphorylated) and inactive (phosphorylated) forms was measured in rat liver mitochondria respiring in state 3 with pyruvate plus malate and 2-oxoglutarate plus malate and during state 4 to state 3 transition in the presence of different substrates. The content of intramitochondrial adenine nucleotides was determined in the parallel experiments. 2. Decrease of the intramitochondrial ATP/ADP ratio with propionate and its increase with palmitoyl-L-carnitine in state 3 is accompanied by a shift of the steady-state of the pyruvate dehydrogenase system towards the active or the inactive form, respectively. 3. Transition from the high energy state (state4) to the active respiration (state3) in mitochondria oxidizing 2-oxoglutarate or plamitoyl-L-carnitine causes an increase of the amount of the active form of pyruvate dehydrogenase due to the decrease of ATP/ADP ratio in the matrix. 4. No change in ATP/ADP ratio can be observed in the presence of octanoate in mitochondria oxidizing pyruvate or 2-oxoglutarate in state 3 or during state 4 to state 3 transition. Simultanelusly, no significant change in phosphorylation state of pyruvate dehydrogenase occurs and a low amount of the enzyme in the active form is present with octanoate or octanoate plus 2-oxoglutarate. Pyruvate abolishes this effect of octanoate and shifts the steady-state of pyruvate dehydrogenase system towards the active form. 5. These results indicate that fatty acids influence the interconversion of pyruvate dehydrogenase mainly by changing intramitochondrial ATP/ADP ratio. However, the comparison of the steady-state level of the pyruvate dehydrogenase system in the presence of different substrates in various metabolic conditions provides some evidence that accumulation of acetyl-CoA and high level of NADH may promote the phosphorylation of pyruvate dehydrogenase. 6. Pyruvate exerts its protective effect against phosphorylation of pyruvate dehydrogenase in the presence of fatty acids of short, medium or long chain in a manner which depends on its concentration. It is suggested that in isolated mitochondria pyruvate counteracts the effect of acetyl-CoA and NADH on pyruvate dehydrogenase kinase.

Adenosine Diphosphate↗
PubMed1994

Mechanisms by which dichloroacetate lowers lactic acid levels: the kinetic interrelationships between lactate, pyruvate, alanine, and glucose.

Dichloroacetate (DCA) is gaining use as an alternative to bicarbonate therapy in the treatment of lactic acidosis. To determine the mechanism(s) by which DCA lowers blood lactate levels, we studied its effect on the kinetic interrelationships between pyruvate, lactate, alanine, and glucose in the hindlimb of dogs during hormonal stimulation of pyruvate production (Ra) and its conversion to lactate. Three groups of dogs (n = 6) were infused with 1-13C-pyruvate to measure whole body pyruvate Ra, and pyruvate Ra and utilization (Rd) across the hindlimb during either a 4-hr infusion of saline (controls), or somatostatin, glucagon, and epinephrine (SGE), or SGE plus dichloroacetate (SGE + DCA). Pyruvate Ra was used as an index of rate of glycolysis and Rd as an index of pyruvate oxidation. In the controls, all kinetic parameters were constant during the saline infusion. Hindlimb pyruvate Ra and Rd were almost equal, and lactate release negligible. Compared to controls, SGE administration significantly increased (P < 0.05) wholebody pyruvate Ra (48.5 +/- 6.2 vs 33.6 +/- 2.4 mumol/kg/min) and blood lactate levels (P < 0.05). Hindlimb pyruvate Ra increased by approximately 150%, but Rd remained unchanged resulting in marked increases in lactate and alanine effluxes. Adding DCA to the SGE infusion significantly reduced wholebody pyruvate Ra (P < 0.05) and blood lactate levels (P < 0.01). In the hindlimb, however, there was no decrease in lactate output, despite a 91% increase in pyruvate utilization because pyruvate Ra also increased. These results suggest that during stimulation of rate of glycolysis, DCA lowers lactate levels by reducing the overall availability of pyruvate for lactate synthesis. This is accomplished by suppressing the rate of glycolysis in tissues other than skeletal muscle and stimulating pyruvate oxidation.

Alanine↗
PubMed1989

Pyruvate metabolism by lymphocytes: evidence for an additional ketogenic tissue.

The rate of utilization of pyruvate (at various concentrations) was measured in lymphocytes prepared from rat mesenteric lymph nodes. The quantitative contribution of pyruvate to CO2, lactate, aspartate, alanine, citrate, acetate, acetyl-CoA and ketone bodies accounted for the pyruvate metabolized. Pyruvate utilization was depressed by increasing concentrations of pyruvate. The maximum catalytic activities and selected intracellular distributions of the following enzymes of pyruvate, citrate and acetyl-CoA metabolism were measured: citrate synthase, ATP-citrate lyase, lactate dehydrogenase, acetyl-CoA hydrolase, acetylcarnitine transferase, NAD+- and NADP+- isocitrate dehydrogenases, HMG-CoA lyase, HMG-CoA synthase, Pyruvate dehydrogenase, acetoacetyl-CoA thiolase, 3-oxoacid-CoA transferase, 3-hydroxybutyrate dehydrogenase and pyruvate carboxylase. Acetyl-CoA formed from pyruvate did not contribute to the respiratory energy metabolism of resting lymphocytes. Instead acetyl-CoA was converted to acetoacetate by reactions which may favour the pathway catalyzed by acetoacetyl-CoA thiolase and 3-oxoacid-CoA transferase. Acetate, acetyl- and palmitoyl-carnitine inhibited the decarboxylation of [1-14C] pyruvate. These observations may be connected with the suppression of pyruvate utilization by increased pyruvate substrate concentration. Only very small amounts of either pyruvate or acetate were incorporated into lipids in resting lymphocytes. The amounts incorporated were partitioned in approximately the same pattern into FFA, T.G., cholesterol and cholesterol esters. Taken together the data show that pyruvate metabolism is directed inter alia at the formation of acetoacetate which may serve as a lipid synthesis precursor. When pyruvate utilization and metabolism was enhanced by concanavalin A, then acetoacetate formation was not favoured and from this it is proposed that the acetyl units may then be directed into lipid synthesis and may also make a contribution to the energy metabolism of the activated lymphocyte.

Animals↗
PubMed1985

The effects of cyclopropane carboxylate on hepatic pyruvate metabolism.

The effects of cyclopropane carboxylate on gluconeogenesis and pyruvate decarboxylation from [1-14C]-labeled pyruvate and lactate were investigated in perfused livers from fasted rats. With high concentrations of pyruvate (greater than or equal to 0.5 mM) in the perfusion medium, infusion of cyclopropane carboxylate inhibited pyruvate decarboxylation and gluconeogenesis by 30 and 40%, respectively. With low, more physiological concentrations of pyruvate (50 microM) or with lactate (1 mM), cyclopropane carboxylate, at a concentration which elicits maximal inhibition of pyruvate decarboxylation from pyruvate (greater than or equal to 0.5 mM), did not affect either pyruvate decarboxylation or gluconeogenesis. Evidence is presented for the rapid formation of the coenzyme-A ester of cyclopropane carboxylate in perfused livers. Infusion of l-(-)carnitine (20 mM) prevented the inhibitory effects of cyclopropane carboxylate on pyruvate decarboxylation and gluconeogenesis from pyruvate (greater than or equal to 0.5 mM). Interestingly, no decrease in the tissue level of cyclopropanecarboxyl-CoA occurs under these conditions. The present study suggests that cyclopropane carboxylate, through a presently ill-defined mediator, inhibits pyruvate decarboxylation and gluconeogenesis by interfering with the pyruvate----oxalacetate----phosphoenolpyruvate----pyruvate cycle when pyruvate (greater than or equal to 0.5mM) supports gluconeogenesis.

Acyl Coenzyme A↗
PubMed1983

Effects of physiologic concentrations of lactate, pyruvate and ascorbate on glucose metabolism in unstressed and oxidatively stressed human red blood cells.

Glucose metabolism was studied in human red blood cells incubated in the presence of physiologic concentrations of ascorbate (0.1 mM) and/or lactate (2 mM) plus pyruvate (0.1 mM). The total flux through glycolysis, as measured by 14C-labeling of glycolytic intermediates, was increased about 15% by ascorbate, 30% by lactate plus pyruvate, and 40% by ascorbate plus lactate plus pyruvate. We found, however, that physiologic concentrations of ascorbate and/or lactate plus pyruvate had no effect on flux of glucose or recycling of pentoses through the hexose monophosphate shunt. Increased formation of lactate accounted for most of the observed increase in glycolysis with little change in pyruvate formation, indicating that the increased flux of reducing equivalents from glucose was stored as lactate rather than being consumed by red cell metabolism. In all experiments, there was a net increase with time in the absolute amount of both lactate and pyruvate in red cell suspensions, indicating that lactate or pyruvate present at zero time did not function as a stoichiometric source or sink for reducing equivalents. There was little effect on steady-state levels of ATP or 2,3-diphosphoglycerate. Equilibration of ascorbate between red cells and the medium was complete before the addition of 14C-labeled glucose to the medium. Glucose metabolism prevented net oxidation of ascorbate in the incubation medium. Physiologic concentrations of ascorbate, lactate and pyruvate appear to increase flux through glycolysis by increasing the turnover of ATP and/or 2,3-diphosphoglycerate. Red cells were exposed to mild oxidative stress by incubation with 0.27 mM 6-hydroxydopamine, 0.27 mM 6-aminodopamine, 0.13 mM 1,4-naphthoquinone-2-sulfonic acid or 0.27 mM phenylhydrazine. The metabolic response to oxidative stress was determined by measuring the formation of methemoglobin, pyruvate, lactate and CO2 in the presence and absence of physiologic concentrations of lactate, pyruvate and ascorbate. Lactate, pyruvate and ascorbate had no effect on the net methemoglobin accumulation but rather on the distribution of the metabolic sources of reducing equivalents and on the flux of reducing equivalents to oxygen. Physiologic lactate and pyruvate allowed increased flow of reducing equivalents from glycolysis to methemoglobin and ultimately oxygen without the necessity of increased flux through glycolysis. This was accomplished by a decrease in the ratio of newly formed lactate to newly formed pyruvate with no increase in total lactate plus pyruvate.(ABSTRACT TRUNCATED AT 400 WORDS)

Adult↗
PubMedDate not supplied

Metabolic control analysis of eucaryotic pyruvate dehydrogenase multienzyme complex.

Metabolic control analysis (MCA) of pyruvate dehydrogenase multienzyme (PDH) complex of eucaryotic cells has been carried out using both in vitro and in vivo mechanistic models. Flux control coefficients (FCC) for the sensitivity of pyruvate decarboxylation rate to activities of various PDH complex reactions are determined. FCCs are shown to be strong functions of both pyruvate levels and various components of PDH complex. With the in vitro model, FCCs are shown to be sensitive to only the E1 component of the PDH complex at low pyruvate concentrations. At high pyruvate concentrations, the control is shared by all of the components, with E1 having a negative influence while the other three components, E2, X, and K, exert a positive control over the pyruvate decarboxylation rate. An unusual behavior of deactivation of the E1 component leading to higher net PDH activity is shown to be linked to the combined effect of protein X acylation and E1 deactivation. The steady-state analysis of the in vivo model reveals multiple steady state behavior of pyruvate metabolism with two stable and one unstable steady-states branches. FCCs also display multiplicity, showing completely different control distribution exerted by pyruvate and PDH components on three branches. At low pyruvate concentrations, pyruvate supply dominates the decarboxylation rate and PDH components do not exert any significant control. Reverse control distribution is observed at high pyruvate concentration. The effect of dilution due to cell growth on pyruvate metabolism is investigated in detail. While pyruvate dilution effects are shown to be negligible under all conditions, significant PDH complex dilution effects are observed under certain conditions. Comparison of in vitro and in vivo models shows that PDH components exert different degrees of control outside and inside the cells. At high pyruvate levels, PDH components are shown to exert a higher degree of control when reactions are taking place inside the cells as compared to the in vitro situation.

Cell Division↗
PubMed2002

Methyl pyruvate stimulates pancreatic beta-cells by a direct effect on KATP channels, and not as a mitochondrial substrate.

In pancreatic beta-cells, methyl pyruvate is a potent secretagogue and is widely used to study stimulus-secretion coupling. In contrast with pyruvate, which barely stimulates insulin secretion, methyl pyruvate was suggested to act as an effective mitochondrial substrate. We show that methyl pyruvate elicited electrical activity in the presence of 0.5 mM glucose, in contrast with pyruvate. Accordingly, methyl pyruvate increased the cytosolic free Ca(2+) concentration after an initial decrease, similar to glucose. The initial decrease was inhibited by thapsigargin, suggesting that methyl pyruvate stimulates ATP production. This assumption is supported by the observation that methyl pyruvate hyperpolarized the mitochondrial membrane potential, similar to glucose. However, in contrast with glucose, methyl pyruvate even slightly decreased NAD(P)H autofluorescence and did not influence ATP production or the ATP/ADP ratio. This observation questions the suggestion that methyl pyruvate acts as a powerful mitochondrial substrate. The finding that methyl pyruvate directly inhibited a cation current across the inner membrane of Jurkat T-lymphocyte mitochondria suggests that this metabolite may increase ATP production in beta-cells by activating the respiratory chains without providing reduction equivalents. We conclude that this mechanism may account for a slight and transient increase in ATP production. We further show that methyl pyruvate inhibited the K(ATP) current measured in the standard whole-cell configuration, an effect that was at least partly antagonized by diazoxide. Accordingly, single-channel currents in inside-out patches were blocked by methyl pyruvate. We conclude that inhibition of K(ATP) channels, and not activation of metabolism, mediates the induction of electrical activity in pancreatic beta-cells by methyl pyruvate.

Adenosine Diphosphate↗
PubMed1978

The effect of fatty acids on the regulation of pyruvate dehydrogenase in perfused rat liver.

The effect of fatty acids on the rate of pyruvate decarboxylation was studied in perfused livers from fed rats. The production of 14CO2 from infused [1-14C]pyruvate was employed as a monitor of the flux through the pyruvate dehydrogenase reaction. A correction for other decarboxylation reactions was made using kinetic analyses. Fatty acid (octanoate or oleate) infusion caused a stimulation of pyruvate decarboxylation at pyruvate concentrations in the perfusate below 1 mM (up to 3-fold at 0.05 mM pyruvate) but decreased the rate to one-third of control rates at pyruvate concentrations near 5 mM. These effects were half-maximal at fatty acid concentrations below 0.1 mM. Infusion of 3-hydroxybutyrate also caused a marked stimulation of pyruvate decarboxylation at low pyruvate concentrations. The data suggest that the mechanism by which fatty acids stimulate the flux through the pyruvate dehydrogenase reaction in perfused liver at low (limiting) pyruvate concentrations involves an acceleration of pyruvate transport into the mitochondrial compartment due to an exchange with acetoacetate. Furthermore, it is proposed that a relationship exists between ketogenesis and the regulation of pyruvate oxidation at pyruvate concentrations approximating conditions in vivo.

Acetoacetates↗