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PubMed2001

Advantages of pyruvate over lactate in peritoneal dialysis solutions.

This review discusses effects of both lactate and pyruvate, and high glucose in peritoneal dialysis solutions (PDS) on leukocytes, mainly on intracellular pH ([pH](i)), glucose metabolic pathways, and apoptosis. Lactate-based PDS (L-PDS) are bioincompatible primarily due to the low pH, high lactate, and glucose excess in both individual and combination. High lactate in an acidi milieu would induce severe intracellular acidosis of leukocytes, and high glucose may disturb glucose metabolic pathways and activate protein kinase C (PKC) and nuclear factor-kappa B (NF-kappaB) of the cells, leading to apoptosis. Pyruvate-based PDS (P-PDS) are novel experimental PDS. Evidence shows that P-PDS are superior in biocompatibility. Pyruvate protection of cells has been confirmed in many fields besides the PDS area. Although the underlying mechanism whereby P-PDS preserve cell function is not fully understood, it may be associated with the maintenance of [pH](i) close to physiological, due to its low buffering capacity, improvement of cellular glucose metabolic pathways and redox state, and sustainment of intracellular calcium ([Ca2+]i) homeostasis in high glucose concentrations. It may also inhibit PKC and NF-kappaB activation in high glucose. In addition, pyruvate is a strong antioxidant, a scavenger of hydrogen peroxide (H2O2). However, exogenous pyruvate in PDS could not be an energy source for cells and also the Crabtree effect might not occur in neutrophils. Pyruvate is a hopeful candidate of buffers in PDS in the near future. Further observation of P-PDS is strongly needed with peritoneal cells to verify the cell protection both in vitro and in vivo before clinic trials.

Apoptosis↗
PubMedDate not supplied

[Effect of pyruvate, threonine, and phosphoethanolamine on acetaldehyde metabolism in rats with toxic liver injury].

Pyruvate dehydrogenase, threonine aldolase and phosphoethanolamine lyase can produce acetaldehyde during normal metabolism. We studied the effect of loading with the substrates of these enzymes (pyruvate, 500 mg/kg, i.p., threonine 500 mg/kg, i.p., and phosphoethanolamine, 230 mg/kg, i.p.) on the blood concentrations of endogenous acetaldehyde and ethanol and the activities of enzymes producing and oxidizing acetaldehyde in the liver of normal rats and rats with liver injury provoked by chronic carbon tetrachloride (CCl4) treatment (0.2 ml i.p. per rat, 2 times a week during 4 weeks). Blood was collected before the treatment and then 30 min and 1 h following the administration of the substrates to intact and CCl4-treated rats. Endogenous acetaldehyde and ethanol were determined by headspace GC. The CCl4 treatment resulted in decreased liver alcohol dehydrogenase and aldehyde dehydrogenase activities and a significant elevation of liver endogenous ehtanol and a clear tendency to enhance blood acetaldehyde levels. Pyruvate increased blood endogenous acetaldehyde in CCl4-treated animals and endogenous ethanol--in the control group of animals. Threonine elevated endogenous acetaldehyde in normal rats. Phosphoethanolamine increased endogenous ethanol in the intact and CCl4 groups. At the same time, in CCl4-treated rats pyruvate administration increased the liver pyruvate dehydrogenase, threonine decreased threonine aldolase, whereas phosphoethanolamine decreased phosphoethanolamine lyase. Thus, the CCl4 effect on blood endogenous acetaldehyde and ethanol may be mediated through decreased liver ALDH and ADH activities. Liver injury promotes the accumulation of acetaldehyde, derived from physiological sources, including the degration of pyruvate and threonine by decreased acetaldehyde oxidation.

Acetaldehyde↗
PubMed2003

Pyruvate improves mitochondrial bioenergetics in an ex-vivo animal model of myocardial ischemia.

Pyruvate is an energy substrate with known cardioprotective activity. We know now that this is due not only to its antioxidant activity, but also to its reduction of intracellular acidosis, modulation of intracytosolic calcium and improvement of cardiomyocyte contractility. However, the role of cardiac mitochondria in such positive effects has only recently begun to be understood and the exact mechanisms of the effect of pyruvate on mitochondria are still largely unknown. Aiming to study the effect of pyruvate on cardiac mitochondrial function during acute ischemia, we used an ex-vivo animal model, perfused in a Langendorff system and then subjected to ischemia in the presence and absence of pyruvate. We evaluated the mitochondrial membrane electrical potential, the respiratory chain O2 consumption (and respiratory control ratio) and the energy charges generated with different energy substrates. We conclude that pyruvate has some effect on the mitochondrial oxidative system (by non-significantly improving the respiratory control ratio), but its main action is on the phosphorylation system, significantly decreasing the time taken to complete a phosphorylation cycle (lag phase) and improving ATP production (increase in energy charge), thus allowing better maintenance of mitochondrial membrane structure, with consequent improvement of the electrical potential after a phosphorylation cycle. These findings have enabled better understanding of the mechanisms behind pyruvate cytoprotection in ischemic cardiomyopathy, clearly highlighting the essential role of cardiac mitochondria in this process.

Animals↗
PubMed1975

Effect of pyruvate and oleate on respiration of frog skeletal and heart muscle.

The authors studied the effect of pyruvate and oleate on O2 consumption of the frog sartorius and heart ventricle. 10 mM pyruvate raised O2 consumption of both tissues by over 100%, but only in the winter. Raised O2 consumption was only partly associated with glycogen synthesis from pyruvate. 0.1 mM oleate reduced O2 consumption in both tissues. A marked drop in O2 consumption was observed in the ventricle (up to 50%). Lecithin had a similar effect on O2 consumption. The addition of pyruvate plus oleate led to an 8-fold increase in O2 consumption of the ventricle, i.e. to maximum oxidation capacity of the tissue, but the addition of lecithin inhibited the pyruvate-induced increase in O2 consumption. It is assumed that both pyruvate and oleate influence resting metabolism in a specific manner which cannot be attributed solely to raised availability of substrate for resting energy metabolism requirements.

Animals↗
PubMed1976

Pyruvate metabolism in rat brain mitochondria.

1. Oxidation of pyruvate by rat brain mitochondria was stimulated in state 3 by malate or succinate up to 250 nmoles O2/mg protein/min. Oxidation of malate, succinate, 2-oxoglutarate or glutamate as the sole substrates, was 1/4 - 1/5 that observed with pyruvate. 2. Maximum oxygen consumption in state 3 was observed at pH 6.90 - 7.20, whereas in state 4 it was not affected by changes in pH. 3. In state 4, in the absence of exogenous acceptor or acetyl residues, acetate was the main oxidation product, corresponding to about 80% of the amount of pyruvate utilized. Malate did not affect the rate of pyruvate utilization but lowered acetate concentration and raised concentration of citrate and 2-oxoglutarate. 4. In state 3, pyruvate and malate were converted mainly to 2-oxoglutarate, its concentration being three times as high as that of citrate. 5. Formation of citrate, 2-oxoglutarate and acetate from pyruvate in brain is considered as a function of availability of the acceptor of acetyl residues and the energy state of mitochondrion.

Adenosine Triphosphate↗
PubMed2004

Ethyl pyruvate: a novel treatment for sepsis and shock.

Pyruvic acid is a simple 3 carbon a-keto-monocarboxylic acid. Recognition that pyruvate is an effective scavenger of reactive oxygen species (ROS) prompted investigators to use it as therapeutic agent for various pathological conditions that are thought to be mediated by redox dependent phenomena, like myocardial, intestinal or hepatic ischemia/reperfusion-induced injury. Ethyl Pyruvate showed to be more effective and safer than equimolar doses of sodium pyruvate. Ethyl Pyruvate showed to have anti-inflammatory effects. In animal models Ethyl Pyruvate improved hyperpermeability and bacterial translocation due to endotoxemia and improved the development of renal disfunction as well as some of the morphological findings of kidney injury. The pharmacological basis for the anti-inflammatory effects of EP remains to be explained. It is plausible that EP mediates suppression of NF-KB activation and secretion of NO and of pro-inflammatory cytokines.

Humans↗
PubMed1992

Double role for pyruvate kinase type M2 in the expansion of phosphometabolite pools found in tumor cells.

As a common characteristic of tumor cells, as well as of normal proliferating cells in the G1-phase of cell cycle, one finds constitutive high levels of all the glycolytic metabolites arising between glucose 6-phosphate and phosphoenolpyruvate. Thus, it is that the phosphometabolites fructose 1,6-bisphosphate, ribose 5-P, P-ribose-PP, NAD, GTP, CTO, UTP, UDP-glucose, glycerol 3-P, glycerol phosphocholine and glycerol phosphoethanolamine are useful in the 31P-nuclear magnetic resonance (NMR) detection of solid tumors in animals and man. This expansion of phosphometabolites is achieved during tumor formation as a result of reductions in levels of enzymes degrading phosphometabolites, owing to the decline in the glycerol 3-P hydrogen shuttle, and as a consequence of alterations in the glycolytic isoenzyme equipment. Tumor cells typically express a particular isoenzyme of pyruvate kinase called type M2 (K) at high levels. This isoenzyme is subject to a complex regulation by amino acids, by fructose 1,6-bisphosphate, and by hormonal- and oncogene-dependent phosphorylation. Pyruvate kinase type M2 is a substrate for the oncogene encoded PP60v-src-tyrosine kinase. A drastic decrease in the affinity for its substrate phosphoenolpyruvate found after transformation by the src-oncogene can be explained as a consequence of the phosphorylation of pyruvate kinase in serine and tyrosine. These phosphorylations induce the breakdown of tetrameric pyruvate kinase to the trimeric and dimeric forms. Unlike the tetrameric form, the dimeric form as a low affinity for phosphoenolpyruvate. Partial inactivation of pyruvate kinase and enolase on the one hand, and a hyperactivation of hexokinase and phosphofructokinase on the other hand, lead to an expansion of all metabolites. Only when these metabolites attain high levels, thereby assuring a sufficient supply of metabolites for RNA, DNA, lipid, and complex carbohydrate synthesis, can cell proliferation proceed. This accumulation of metabolites in the G1-phase cells has been termed a "metabolic budget system" because it senses not only the actual nutrient levels, but also the supply over a period of time. Monoclonal antibodies specific for the dimeric form of pyruvate kinase type M2 can be used for the immunohistological detection of tumor cells. The amount of the dimeric form in tumor cells closely correlates with the degree of malignancy and can be used for a nonspecific detection of tumors based on assays performed with patient's plasma.

Adenosine Triphosphate↗
PubMed2003

[CaCO3 stimulates alpha-ketoglutarate accumulation during pyruvate fermentation by Torulopsis glabrata].

A large amount of alpha-ketoglutarate (alpha-KG) (6.8 g/L) was accumulated in flask culture when CaCO3 was used as a buffering agent in the production of pyruvate by multi-vitamin auxotrophic yeast Torulopsis glabrata CCTCC M202019. In a 5 L jar-fermentor, less alpha-KG (1.3 g/L) was produced when NaOH was used to adjust the pH, while more alpha-KG (11.5 g/L) detected when CaCO3 was used as the buffer. In the latter case, the molar carbon ratio of pyruvate to alpha-KG (C(PYR)/ CalphaKG) was similar to that obtained in flask culture, suggesting the accumulation of alpha-ketoglutarate was related to the addition of CaCO3. Furthermore, it was found that: (1) delaying the addition time of CaCO3 decreased the a-ketoglutarate formation but increased C(PYR)/ C(alphaKG); and (2) under vitamin limitation conditions increasing the concentration of CaCO3 led to an increased a-KG accumulation at the expenses of pyruvate. To study which ions in CaCO3 was responsible for the accumulation of alpha-KG, the effects of different pH buffers on the a-KG accumulation were studied. The level of alpha-KG was found to correlate with the levels of both Ca2+ and CO3(2-), with Ca2+ played a dominant role and CO3(2-) played a minor role. To find out which pathway was responsible for the accumulation of alpha-KG, the effects of biotin and thiamine on alpha-KG accumulation was investigated. The increase in biotin concentration led to an increase in alpha-KG accumulation and a decrease in C(PYR)/ C(alpha-KG), while the levels of alpha-KG and C(PYR)/C(alphaKG) were not affected by thiamine concentration. The activity of pyruvate carboxylase was increased as much as 40% when the medium was supplemented with Ca2+ . On the other hand, the activity of the pyruvate dehydrogenase complex was unaffected by the presence of Ca2+. To conclude, the higher level of a-KG was caused by higher activity of pyruvate carboxylase stimulated by Ca2+, with CO3(2-) served as the substrate of the reaction.

Biotechnology↗
PubMed2005

The force-frequency relationship in human heart failure: effect of pyruvate and isoproterenol.

PURPOSE: The purpose of present study was to investigate the effect of metabolic substrate pyruvate and beta-adrenergic agonist isoproterenol and combination of these agents on the force- and relaxation-frequency relationship in human heart failure. MATERIAL AND METHODS: The experiments were performed on isolated human ventricle strips from patients undergoing cardiac corrective open heart surgery, using conventional method of registration of electromechanical activity. The stimulation frequency of myocardial strips was 0.2, 0.5, 1.0, 1.5, 2.0, 2.5 and 3.0 Hz. RESULTS: In control, i.e. at perfusion of myocardial strips by Tyrode solution and stimulation frequency 1 Hz, the contraction force (F) was 0.94 +/- 0.18 mN, half time of relaxation (tr)--178.8 +/- 9.3 ms (n= 12). Pyruvate (10 mmol/L) increased F to 176.0 +/- 13.4%, tr--104.6 +/- 3.1% (n=8, p<0.05) vs control. By the action of isoproterenol (10(-5) mol/L) F increased to 122.1 +/- 10.2%, tr decreased to 58.9 +/- 3.1% (n=4, p<0.05) vs control. The relationship of F and tr from stimulation frequency in the absence of pyruvate and isoproterenol was negative. Pyruvate and isoproterenol didn't alter the shape of force-frequency relationships but F was augmented at all stimulation frequencies. The positive inotropic effect of isoproterenol was potentiated by pyruvate. CONCLUSIONS: Pyruvate and isoproterenol alone can improve cardiac contractility in wide-range of stimulation frequency. The combination of these inotropic agents results in even more effective increase of contractile performance and therefore may be of therapeutic value in heart failure.

Adrenergic beta-Agonists↗
PubMed2006

[Effects of temperature on the kinetics and level of energy charge and oxidation-reduction state in pyruvate biosynthesis].

Temperature plays an important role in pyruvate biosynthesis by Torulopsis glabrata 620. The effects of temperature on the substrate consumption, cell growth, pyruvate biosynthesis and level of energy charge and oxidation-reduction state have been investigated. During the constant temperature fermentation, higher temperature can enhance the rate of glucose consumption, cell growth and pyruvate production. However, it also leads to higher energy charge in the prophase of fermentation, too much consumption of glucose by cell and deficient throughput of pyruvate in the anaphase of fermentation, which brings on decrease of pyruvate yield. Oppositely, maintaining lower temperature during the fermentation can offer constant production capacity of pyruvate in the anaphase of fermentation. But the disadvantages are lower cell growth and higher level of NADH/NAD+ in vivo during the anaphase of fermentation, which leads to lower productivity inevitably. Disaccords above-mentioned indicate that the higher production, higher yield and higher productivity cannot be achieved at one time during a constant temperature fermentation.

Candida glabrata↗
PubMed1979

[Conformational isomerization of lactate dehydrogenase complexes formed by pyruvate and coenzyme analogs].

The kinetics of LDH-catalyzed reduction of pyruvate involving APADH were studied. It was shown that under conditions of a single turnover reaction the first order rate constant is equal to 37+/-4 sec-1. The reaction rate (vo) did not change when a deutero-coenzyme was used. The relationship between vo and pyruvate concentration is hyperbolic. It is concluded that isomerization of the ternary LDH-APADH-pyruvate complex limits the reaction rate. The spectral properties and the kinetics of formation and dissociation of abortive LDH complexes with pyruvate and NAD analogs (APAD and PAAD) were studied. The participation of the carboxamide group of NAD in conformational isomerization of the LDH-NADH-pyruvate and LDH-NAD-pyruvate complexes was studied.

Catalysis↗
PubMed1989

Expression of the rat L-type pyruvate kinase gene from its dual erythroid- and liver-specific promoter in transgenic mice.

The gene for the L-type pyruvate kinase possesses two promoters which are located 500 base pairs apart. The L promoter is specific to liver and regulated by hormones and diet; the L' promoter is specific to erythroid cells. We produced two series of transgenic mice carrying either the entire rat L-pyruvate kinase gene or a minigene devoid of exons two to nine, with 2.7 kilobases of flanking sequences 5' to the cap site of the L' promoter and 1.4 kilobases 3' to the downstream polyadenylation site. In both series the patterns of expression from the two promoters were similar to those of the endogenous rat gene. The rat L promoter was expressed strongly in liver and weakly in kidney and gut of adult transgenic mice. Moreover, it was regulated like the endogenous rat L-pyruvate kinase gene upon hormonal and nutritional adaptation: the level of L-pyruvate kinase mRNA was decreased dramatically by 24 h of starvation, while refeeding a carbohydrate-rich diet strongly stimulated expression of the transgenes. This stimulation was prevented by glucagon. Use of alternative polyadenylation sites in the last exon of the rat L-type pyruvate kinase gene was similar for both types of transgenes and similar to that in rat and not control mice, suggesting that the transgenes contain the sequences that control the choice of polyadenylation site. Transcription of the minigene was higher than that of the entire transgene, probably due to the high copy number of the minigene. At the protein level, rat L subunits encoded by the entire transgene were more abundant than mouse subunits in the liver of adult transgenic mice. In contrast, expression of the rat L' promoter in fetal liver was only 5% of that in fetal rat liver, and we were unable to detect rat L' subunits of pyruvate kinase enzyme in the red blood cells from transgenic mice. Our results suggest that the integrated DNA contains all elements necessary for tissue specificity (L' and L) as well as hormonal and nutritional control (L) of expression of the rat transgene. Nevertheless, a L'-specific activating element may be missing.

Animals↗
PubMed1989

Metabolic effects of high glucose concentrations: inhibition of hepatic pyruvate kinase.

We tested the in vitro effects of various glucose concentrations on the activity of hepatic pyruvate kinase, assayed at subsaturating, near physiological concentration (0.20 mmol/l) of the substrate phosphoenolpyruvate, to detect the "active" form of the enzyme. A 10-min incubation of mouse liver slices (n = 18) with increasing glucose concentrations (5, 10 and 20 mmol/l) resulted in a significant (p less than 0.01), progressive pyruvate kinase inhibition of 15, 28 and 41%, respectively. Similar data were obtained by incubating mouse liver homogenates (n = 7) with glucose, although with this material (which was supplemented with the pyruvate kinase activator fructose-1,6-diphosphate) the inhibition at the highest glucose concentration used was lower (24%, p less than 0.02). Addition of 10 nmol/l insulin during slice incubation (n = 8) prevented by 98% and 69% the inhibition exerted by 10 and 20 mmol/l glucose, respectively. Insulin alone was without effect on the enzyme activity. Glucose might inhibit pyruvate kinase by competing with the activator fructose-1,6-diphosphate. Insulin might overcome the glucose effect by activating pyruvate kinase through the known mechanism of enzyme dephosphorylation. Thus, in decompensated diabetes the high level of blood glucose may contribute, together with the counterregulatory hormones, to inhibit hepatic pyruvate kinase and therefore to stimulate gluconeogenesis.

Animals↗
PubMed1988

Primary structure of maize pyruvate, orthophosphate dikinase as deduced from cDNA sequence.

We have isolated two overlapping cDNA clones that encompass the entire structural gene for pyruvate, orthophosphate dikinase from maize. The analysis of the nucleotide sequence has revealed that the cDNA clones include an insert of a total of 3,171 nucleotides without a poly(A) tail and encode a polypeptide that contains 947 amino acid residues and has a molecular weight of 102,673. Comparison of the N-terminal amino acid sequence of purified pyruvate, orthophosphate dikinase protein with that deduced from the nucleotide sequence shows that the mature form of pyruvate, orthophosphate dikinase in the maize chloroplast consists of 876 amino acid residues and has a molecular weight of 95,353. The amino acid composition of the deduced sequence of pyruvate, orthophosphate dikinase is in good agreement with that of the purified enzyme. The region that contains the active and regulatory sites of pyruvate, orthophosphate dikinase can be found in the deduced sequence of amino acids. We have predicted the secondary structure and calculated the hydropathy pattern of this region. The extra 71 residues at the N terminus of the deduced sequence of amino acid residues corresponds to the transit peptide which is indispensable for the transport of the precursor protein into chloroplasts. We have compared the primary structure of the pyruvate, orthophosphate dikinase transit peptide to those of other proteins and found sequences similar to the consensus sequences found in other transit peptides.

Amino Acid Sequence↗
PubMedDate not supplied

[Effect of anions on inhibition of lactate dehydrogenase by pyruvate].

The effect of anions Cl- and I- on structural and kinetic properties of LDH was investigated. It was shown that anions are specific inhibitors of LDH competing with pyruvate in the active ternary complex, LDHNADHpyq. The following dissociation constants for the anions were obtained from inhibition data: 0.4 +/- 0.02 and 0.07 +/- 0.01 M for Cl- and I-, respectively. The slope of Hill plot are near 1.0. The anions abolished the inhibition of LDH at high pyruvate concentrations. The following dissociation constants were obtained from these data: 0.1 and 0.015 M for Cl- and I- respectively. The inhibition by anions and the abolishing of substrate inhibition by anions were studied also for the lactate oxidation reaction. The dissociation constants for anions obtained from these data are in good correlation with the constants obtained for the pyruvate reduction reaction. It was concluded that anions do not interact with the group at the catalytic site with pK approximately 7.8, presumably His-195. The degree of pyruvate inhibition does not depend on the buffer system. The differences in the degree of inhibition obtained previously in phosphate, imidazole and tris-buffer systems can be explained by the presence of Cl- anions in the last two buffer. The rate constants of hydroxy leads to keto pyruvate transition was obtained in various buffer systems. It was shown that the hydroxy-form of pyruvate does not cause the inhibition of LDH.

Animals↗
PubMed1988

Sulfonylureas inhibit metabolic flux through rat liver pyruvate carboxylase reaction.

The effect of oral hypoglycemic sulfonylureas, tolbutamide and glyburide, on metabolic flux through the pyruvate carboxylase reaction was evaluated in liver mitochondria isolated from 24-hr fasted rats. Both these sulfonylureas inhibited the metabolic flux through the pyruvate carboxylase reaction in a concentration dependent manner. Half-maximal inhibition was achieved at tolbutamide and glyburide concentrations of 0.85 mM and 63.3 microM, respectively. Neither sulfonylurea altered the activity of pyruvate carboxylase or the Km of the enzyme for ATP and pyruvate. However, glyburide and tolbutamide decreased mitochondrial ATP content and elevated mitochondrial ADP and AMP levels. The decrease in mitochondrial ATP was greater with 400 microM glyburide compared with 2.0 mM tolbutamide. Glyburide also decreased mitochondrial acetyl-coenzyme A/CoASH ratio. Additionally, glyburide and tolbutamide stimulated pyruvate (5 mM) supported mitochondrial respiration in the absence of ADP. These data indicate that these sulfonylureas inhibit the metabolic flux through the pyruvate carboxylase reaction by decreasing mitochondrial ATP/ADP and acetyl-coenzyme A/CoASH ratios. Decreased mitochondrial nucleotide content and increased mitochondrial respiration caused by sulfonylureas suggest that these compounds may uncouple oxidative phosphorylation.

Acetyl Coenzyme A↗
PubMed1987

Carbon isotope effects on the pyruvate dehydrogenase reaction and their importance for relative carbon-13 depletion in lipids.

A method has been developed for the positional 13C isotope analysis of pyruvate and acetate by stepwise quantitative degradation. On its base, the kinetic isotope effects on the pyruvate dehydrogenase reaction (enzymes from Escherichia coli and Saccharomyces cerevisiae) for both of the carbon atoms involved in the bond scission (double isotope effect determination) and on C-3 of pyruvate have been determined. The experimental k12/k13 values with the enzyme from E. coli on C-1 and C-2 of pyruvate are 1.0093 +/- 0.0007 and 1.0213 +/- 0.0017, respectively, and, with the enzyme from S. cerevisiae, the values are 1.0238 +/- 0.0013 and 1.0254 +/- 0.0016, respectively. A secondary isotope effect of 1.0031 +/- 0.0009 on C-3 (CH3-group) was found with both enzymes. The size of the isotope on C-1 indicates that decarboxylation is more rate-determining with the yeast enzyme than with the enzyme from E. coli, although it is not the entirely rate-limiting step in the overall reaction sequence. Assuming appropriate values for the intrinsic isotope effect on the decarboxylation step (k3) and the equilibrium isotope effect on the reversible substrate binding (k1, k2), one can calculate values for the partitioning factor R (k3/k2: E. coli enzyme 4.67, S. cerevisiae enzyme 1.14) and the intrinsic isotope effects related to the carbonyl-C (k1/k'1 = 1.019; k3/k'3 = 1.033). The isotope fractionation at C-2 of pyruvate gives strong evidence that the well known relative carbon-13 depletion in lipids from biological material is mainly caused by the isotope effect on the pyruvate dehydrogenase reaction. In addition, our results indicate an alternating 13C abundance in fatty acids, that has already been verified in some cases.

Carbon Isotopes↗
PubMedDate not supplied

Characterization of a pyruvate dehydrogenase modulator purified from insulin-treated rat brain plasma membranes.

A factor able to stimulate pyruvate dehydrogenase when added to purified mitochondria was prepared from the supernatant of brain plasma membranes incubated with physiological concentrations of insulin (25 microU/ml). The factor completely reactivated pyruvate dehydrogenase previously inhibited with ATP and was active on pyruvate dehydrogenase from brain and liver mitochondria and from peripheral lymphocytes. The insulin-dependent stimulator of pyruvate dehydrogenase was heat and acid stable, was not absorbed on charcoal and displayed an isoelectric point of 5.5. The insulin mediator was purified by gel filtration, DEAE-cellulose and sulfonated polystyrene chromatography and, after dansylation, by high performance liquid chromatography. The purified mediator displayed a molecular weight of about 2800 and appeared as a peptide rich in glycine and serine and void of proline and sulfur containing aminoacids. It retained its stimulatory action on pyruvate dehydrogenase after dansylation and was completely inactivated by trypsin and chymotrypsin. Full reactivation of ATP-inhibited pyruvate dehydrogenase was attained when mitochondria were incubated with a mediator concentration of about 0.5 microM.

Amino Acids↗