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PubMed2003

Presence of D-alanine in an endopeptidase from Streptococcus pyogenes.

D-amino acids are commonly found in peptide antibiotics and the cell wall peptidoglycan of bacterial cell walls but have not been identified in proteins or enzymes. Here we report the presence of 6-7 A-alanine residues in an endopeptidase of Streptococcus pyogenes, a unique enzyme involved in surface protein attachment that we term LPXTGase. Using D-amino acid oxidase coupled with catalase for the deamination of D-alanine to pyruvic acid (a conversion unique to D-alanine), we were able to identify [14C]pyruvic acid in a [14C]alanine-labeled preparation of purified LPXTGase, which represents 27% of the amino acid composition. Because D-amino acids are not accommodated in ribosomal peptide synthesis, these results suggest that the same process used in assembling peptide antibiotics or a yet unidentified mechanism may synthesize the core protein of this endopeptidase.

Alanine↗
PubMed1967

Intracellular localization of nitrate reductase, nitrite reductase, and glutamic Acid dehydrogenase in green leaf tissue.

Greenhouse grown seedlings of corn (Zea mays L.) and foxtail (Setaria faberii Herrm.) were used as source material in determining the intracellular localization of nitrate reductase, nitrite reductase, and glutamic acid dehydrogenase, Nonaqueous and aqueous isolation techniques were used to establish that nitrite reductase is localized within the chloroplasts, but that nitrate reductase and glutamic acid dehydrogenase are not. Nonaqueous isolation gives distribution patterns of nitrite reductase which are the same as those observed for NADP-dependent 3-phosphoglyceraldehyde dehydrogenase but which differ drastically from the patterns observed for pyruvic acid kinase. The distribution patterns for nitrate reductase are the same as those of pyruvic acid kinase. The techniques used do not eliminate the possibility that nitrate reductase and pyruvic acid kinase are localized on the external chloroplast membrane.The data obtained establish that glutamic acid dehydrogenase of green leaves is localized within the mitochondria.

Journal Article↗
PubMed1970

Degradation of a pneumococcal type-specific polysaccharide with exposure of group-specificity.

Pyruvic acid is an immunological determinant of the quite rigorously type-specific capsular polysaccharide of pneumococcal type IV (S IV). Removal of pyruvic acid by mild hydrolysis converts the capsular polysaccharide of type IV into an analog of the pneumococcal group-specific C-substance. Depyruvylated S IV resembles C-substance so closely immunologically that it not only precipitates a high proportion of the anti-C in antipneumococcal sera, regardless of their immunological types, but also, like C, precipitates human C-reactive protein in the presence of calcium ions. Apparently, the removal of pyruvic acid ketal rings from adjacent sugars unmasks N-acetylgalactosamine residues which must be linked and spaced much as are those in C-substance. Groupings reactive with suitably linked N-acetylgalactosamine, therefore, appear to be located on the surfaces of molecules of human C-reactive protein.

Antigen-Antibody Reactions↗
PubMedDate not supplied

Utilization of an alternative carbon source for efficient production of human alpha(1)-antitrypsin by genetically engineered rice cell culture.

Human alpha(1)-antitrypsin was produced by genetically engineered rice cells using promoter and signal peptide of a rice alpha-amylase isozyme. Batch and continuous cultures were employed to investigate the effects of alternative carbon sources on the alpha(1)-antitrypsin production. While this expression system is inducible by sugar depletion, we have found that the productivity of alpha(1)-antitrypsin increased 2.4- to 3.4-fold, compared with the control medium without carbon source, in medium containing an alternative carbon source, such as pyruvic acid and glyoxylic acid. The accumulated alpha(1)-antitrypsin in the medium containing pyruvic acid reached 18.2-24.2 mg/g-dry cell in 50-70 h by batch culture.

Carbon↗
PubMed1996

Cloning and characterization of a locus encoding an indolepyruvate decarboxylase involved in indole-3-acetic acid synthesis in Erwinia herbicola.

Erwinia herbicola 299R synthesizes indole-3-acetic acid (IAA) primarily by the indole-3-pyruvic acid pathway. A gene involved in the biosynthesis of IAA was cloned from strain 299R. This gene (ipdC) conferred the synthesis of indole-3-acetaldehyde and tryptophol upon Escherichia coli DH5 alpha in cultures supplemented with L-tryptophan. The deduced amino acid sequence of the gene product has high similarity to that of the indolepyruvate decarboxylase of Enterobacter cloacae. Regions within pyruvate decarboxylases of various fungal and plant species also exhibited considerable homology to portions of this gene. This gene therefore presumably encodes an indolepyruvate decarboxylase (IpdC) which catalyzes the conversion of indole-3-pyruvic acid to indole-3-acetaldehyde. Insertions of Tn3-spice within ipdC abolished the ability of strain 299R to synthesize indole-3-acetaldehyde and tryptophol and reduced its IAA production in tryptophan-supplemented minimal medium by approximately 10-fold, thus providing genetic evidence for the role of the indolepyruvate pathway in IAA synthesis in this strain. An ipdC probe hybridized strongly with the genomic DNA of all E. herbicola strains tested in Southern hybridization studies, suggesting that the indolepyruvate pathway is common in this species. Maximum parsimony analysis revealed that the ipdC gene is highly conserved within this group and that strains of diverse geographic origin were very similar with respect to ipdC.

Amino Acid Sequence↗
PubMed1982

Regulation of the branched chain alpha-keto acid and pyruvate dehydrogenases in the perfused rat heart.

Phosphorylation of the alpha-subunits of the branched chain alpha-keto acid and the pyruvate dehydrogenases was measured in mitochondria isolated from rat hearts perfused in the presence of [32P]phosphate and various substrates. Mitochondrial isolations were accomplished rapidly under conditions which would preclude the interconversion of these two enzyme complexes between their active (dephosphorylated) and inactive (phosphorylated) forms. Inactivation of the branched chain complex and the concomitant incorporation of [32P]phosphate into the alpha-chain of the dehydrogenase subcomponent were observed in hearts perfused with glucose (10 mM) or pyruvate (10 mM). alpha-Ketoisocaproate infusion caused a 15-fold activation and dephosphorylation of the branched chain dehydrogenase, while pyruvate dehydrogenase remained inactive and phosphorylated under these conditions. Both dehydrogenase components were partially activated, and most of the [32P]phosphate was removed from their respective alpha-subunits during substrate-free perfusion. Both enzyme complexes were activated and dephosphorylated completely during infusion of dichloroacetate (1 mM) and glucose (10 mM).

3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗
PubMedDate not supplied

Effect of varicocele on sperm respiration and metabolism.

Semen samples from 41 infertile male patients (30 with varicocele and 11 controls) were examined, lactic and pyruvic acids were measured and the oxygen consumption was determined. Decreased sperm count, diminished motility, decreased lactic acid concentration and oxygen consumption were noticed among the varicocele patients as compared to controls. Pyruvic acid concentration was slightly increased in the varicocele group. It was concluded that a decrease in lactic dehydrogenase activity may be one of the abnormalities in the semen of varicocele patients.

Humans↗
PubMed1993

Xylitol-induced increase in purine degradation: a role of erythrocytes.

We administered xylitol intravenously to normal subjects to investigate the mechanism of xylitol-induced increase in the purine degradation in humans. Xylitol increased the plasma concentrations of hypoxanthine, xanthine and uric acid but decreased the blood concentration of pyruvic acid. The erythrocyte concentrations of IMP, AMP, ADP, glyceraldehyde 3-phosphate and fructose 1,6-diphosphate as well as the urinary excretion of hypoxanthine and xanthine were increased, while the erythrocyte concentration of ATP was decreased. In addition, the in vitro incubation studies using erythrocytes demonstrated that both xylitol-induced purine degradation and xylitol-induced inhibition of the conversion of glyceraldehyde 3-phosphate to 1,3-bisphosphoglycerate were protected by pyruvic acid. These results indicate that xylitol-induced impairment of glycolysis in erythrocytes contributes to the observed xylitol-induced increase in the purine degradation in the body.

Adult↗
PubMed1999

Effects of exposure to low concentrations of carbon monoxide on exercise performance and myocardial perfusion in young healthy men.

OBJECTIVE: To assess the effects of exposure to low concentrations of carbon monoxide (CO), as commonly measured in atmospheric urban air pollution and certain occupational environments, on exercise performance and myocardial perfusion in young healthy men, and the possible need for tighter restrictions on ambient concentrations of CO. METHODS: 15 young, healthy non-smoking men, 18-35 years old, were exposed blindly and randomly to air or to a mixture of CO and air, followed by an exercise treadmill test with thallium heart scintigraphy. Blood was drawn for determination of carboxyhaemoglobin before and at the end of the exposure, and for lactic and pyruvic acid at the beginning and the end of the exercise test. The main outcome measures include the duration of the exercise test, the maximal effort expressed in metabolic equivalent units (METs), the mean plasma lactic to pyruvic acid ratio at the end of the ergometry, ECG changes in the exercise test, and perfusion deficits in thallium heart scintigraphy. RESULTS: At the end of exposure to CO, the mean (SD) blood carboxyhaemoglobin concentration rose from 0.59% (0.08%) to 5.12% (0.65%) (p < 0.0001). At the end of the exercise period, the mean (SD) plasma lactate/pyruvate ratio, which reflects the level of anaerobic metabolism (69.9 (5.9) after air and 75.9 (7.0) after CO), was not significantly different between the two experimental groups. Exercise induced electrocardiographic changes were noted in only one subject after exposure to CO. No arrhythmias were detected in any of the subjects. Significant differences were found in the mean duration of the exercise test (p = 0.0012) and the METs (p = 0.0001). The mean adjusted difference of exercise duration between exposure to air and CO was 1.52 minutes 95% confidence interval (95% CI) 0.73 to 2.32 minutes. The mean adjusted difference of METs between exposure to air and CO was 2.04 95% CI 1.33 to 2.76. The models for duration of exercise and METs showed no significant sequence and period effects. Thallium myocardial perfusion imaging disclosed normal perfusion in all regions of the heart, with no significant differences in perfusion between the two exercise tests (after air or CO). CONCLUSION: Acute exposure to a low concentration of CO which produces blood carboxyhaemoglobin concentrations of 4%-6% significantly decreases exercise performance in young healthy men. No ischaemic electrocardiographic changes or disturbances in myocardial perfusion were found by graded exercise with thallium scintigraphy. Our findings suggest that pollution of atmospheric air by CO at concentrations which are commonly found in urban and industrial environments may exert an adverse effect on skeletal muscles, manifesting as decreased exercise performance.

Adolescent↗
PubMed1988

Determination of glycolytic intermediates in a flow injection system using immobilized enzymes.

Some glycolytic enzymes (lactate dehydrogenase, pyruvate kinase, enolase and phosphoglyceromutase) were immobilized on a polyacrylamide-type bead polymer containing carboxylic functional groups activated by water-soluble carbodiimide. The immobilized enzymes were used for the determination of pyruvic acid, phosphoenolpyruvic acid, 2-phosphoglyceric acid and 3-phosphoglyceric acid in a flow injection system. The immobilized lactate dehydrogenase column was repeatedly employed for the determination of pyruvic acid in clinical samples. The results of the flow injection method accorded well in accuracy, sensitivity and reproducibility with those of soluble enzyme analysis.

Enzyme Stability↗
PubMed2000

[Research of indole-3-acetic acid biosynthetic pathway of Klebsiella oxytoca SG-11 by HPLC and GC-MS].

The plant growth promoting bacteria are closely associated to plant. The bacteria are used to adhering to plant rhizoplane, promoting plant growth by fixing nitrogen from atmosphere, secreting stimulating substances or producing antagonistic to plant pathogens. It was indicated that the biological nitrogen fixation played an important role in plant growth promoting function. In fact, it was verified recently by overall research that IAA does it. Therefore research of IAA production and biosynthetic pathway of plant growth-promoting bacteria is much more important. The various ways of IAA production indicated the strong or weak promoting function of bacterium to plants in general. The purpose of this paper is to determine whether IAA exists in cultured medium of Klebsiella axytoca SG-11 and biosynthetic pathway of IAA, in order to opimize cultural conditions for IAA production. Klebsiella axytoca SG-11 is a plant growth promoting bacterium, isolated from rice rhizoplane, which can fix nitrogen. The supernatant of SG-11 cultured medium determined by HPLC showed that 47.4 mg/L of IAA existed in LB medium and 1.2 mg/L of IAA, in basal medium. IAA in metabolite was identified by GC/MS as well. The intermediate determination of tryptamine, indole-3-acetamide, tryptophol and indole-3-acetonitrile indirectly indicated that IAA was biosynthesized in a pathway of indole-3-pyruvic acid. Meanwhile, tryptophol in metabolite of SG-11 was verified by GC/MS. The direct intermediates of indole-3-pyruvic acid and indole-3-acetaldehyde in the pathway can not be determined, because both are unstable under normal condition. As reversible conversion existed between indole-3 pyruvic aldehyde and tryptophol, the presence of tryptophol also proved the pathway of indole-3-pyruvic acid in the synthesis of IAA by Klebsiella axytoca SG-11. The results laid basis for further research of plant growth-promoting function of the bacterium.

Chromatography, High Pressure Liquid↗
PubMed1982

The interaction of granaticin with nucleic acids and pyruvate decarboxylase.

The interaction of granaticin with two different yeast ribonucleic acids, dialyzed transfer RNA and calf thymus DNA was studied. Contrary to previous reports, no binding of granaticin to DNA or RNA was observed. The visible spectrum of granaticin on direct mixing or equilibrium dialysis of granaticin with RNA or DNA was unchanged. Furthermore, granaticin did not displace acridine orange from DNA in competitive binding studies using fluorescence polarization. However, granaticin was shown to inhibit pyruvate decarboxylase. From the Ki for granaticin (3.8 mM) it was concluded that granaticin is as efficient as other 1,4-naphthoquinones in inhibiting pyruvate decarboxylase.

Animals↗
PubMed1994

Ouabain and digitoxin as modulators of chick embryo cardiomyocyte energy metabolism.

The effects of 0.1 nmol/l ouabain (CAS 630-60-4) and digitoxin (CAS 71-63-6) on oxygen consumption rate (OCR) and the contractility of cultured chick embryo cardiomyocytes were investigated using a perfusion system which allowed microscopic observation during the experiment. contractility was assessed by a novel image analysis procedure. During substrate free perfusion the OCR was increased on application of 0.1 nmol/l ouabain or digitoxin for 60 min, whilst contractility was not affected. Digitoxin (0.1 nmol/l) elicited no change in the OCR in the presence of glucose, pyruvic acid, lactic acid and caprylic acid. Ouabain (0.1 nmol/l) did not affect the OCR or contractility in the presence of glucose, pyruvic acid and lactic acid, however together with caprylic acid the OCR was increased significantly. These results demonstrate a hitherto unknown stimulation of fatty acid utilization by low concentrations of ouabain.

Animals↗
PubMed1984

[Shifts in tissue respiration of the lung and liver in rabbits administered penicillin and streptomycin].

The experiments were performed on intact rabbits weighing 2.5-3 kg. The animals were treated with penicillin or streptomycin injected intramuscularly in doses of 20 000 units or 17 000 micrograms a day per kg bw for 12 days. The concentrations of lactic and pyruvic acids, cytochrome c, activity of cytochrome c oxidase and thiol spectrum were investigated in the lungs and liver immediately after the antibiotic use in 10 rabbits of every experimental series. To follow the period of persisting of the antibiotic-induced changes the other 10 rabbits were killed 29 days after the last injection of the drugs. It was shown that penicillin induced significant changes in the concentrations of lactic and pyruvic acids in the lung tissue, activated cytochrome c oxidase and decreased the concentrations of free SH and S-S groups immediately after its administration. 29 days after discontinuation of penicillin the activity of cytochrome c oxidase returned to normal, while the other biochemical indices remained changed. Penicillin induced changes in the concentrations of lactic and pyruvic acids and the activity of cytochrome c oxidase in the liver. The concentrations of free masked SH and S-S groups also changed. 29 days after discontinuation of penicillin the enzymatic system of cytochrome c-cytochrome c oxidase alone returned to normal. Like penicillin, streptomycin induced changes in the tissue respiration of the lungs and liver. The character of the streptomycin-induced changes was more pronounced both immediately and 29 days after discontinuation of the drug use.

Animals↗
PubMed1982

Effect of the fatty acid oxidation inhibitor 2-tetradecylglycidic acid on pyruvate dehydrogenase complex activity in starved and alloxan-diabetic rats.

Intravenous administration of the fatty acid oxidation inhibitor 2-tetradecylglycidic acid had no effect on the proportion of pyruvate dehydrogenase complex in the active form in heart, diaphragm or gastrocnemius muscles or in liver, kidney or adipose tissue of fed normal rats. The compound reversed the effect of 48h starvation (which decreased the proportion of active complex) in heart muscle, partially reversed the effect of starvation in kidney, but had no effect in the other tissues listed. The compound failed to reverse the effect of alloxan-diabetes (which decreased the proportion of active complex) in any of these tissues. In perfused hearts of fed normal rats, 2-tetradecylglycidate reversed effects of palmitate (which decreased the proportion of active complex), but it had no effect in the absence of palmitate. In perfused hearts of 48h-starved rats the compound increased the proportion of active complex to that found in fed normal rats in the presence or absence of insulin. In perfused hearts of diabetic rats the compound normalized the proportion of active complex in the presence of insulin, but not in its absence. Palmitate reversed the effects of 2-tetradecylglycidate in perfused hearts of starved or diabetic rats. Evidence is given that 2-tetradecylglycidate only reverses effects of starvation and alloxan-diabetes on the proportion of active complex in heart muscle under conditions in which it inhibits fatty acid oxidation. It is concluded that effects of starvation and alloxan-diabetes on the proportion of active complex in heart muscle are dependent on fatty acid oxidation. Insulin had no effect on the proportion of active complex in hearts or diaphragms of fed or starved rats in vitro. In perfused hearts of alloxan-diabetic rats, insulin induced a modest increase in the proportion of active complex in the presence of albumin, but not in its absence.

Acetyl Coenzyme A↗
PubMed1999

Effect of citric acid on the biosynthesis and composition of xanthan.

The effect of citric acid metabolism by Xanthomonas campestris on composition of xanthan has been studied. Citric acid consumption in fed-batch and continuous fermentation increased the pyruvic acid content of xanthan. An increase in pyruvic acid content in xanthan has been explained with the help of energy balance in xanthan biosynthesis.

Journal Article↗
PubMed2003

The role of glycolysis and gluconeogenesis in the cytoprotection of neuroblastoma cells against 1-methyl 4-phenylpyridinium ion toxicity.

1-Methyl-4-phenylpyridinium (MPP+) is a mitochondrial Complex I inhibitor and is frequently used to investigate the pathological degeneration of neurons associated with Parkinson's disease (PD). In vitro, extracellular concentration of glucose is one of the most critical factors in establishing the vulnerability of neurons to MPP+ toxicity. While glucose is the primary energy fuel for the brain, central nervous system (CNS) neurons can also take up and utilize other metabolic intermediates for energy. In this study, we compared various monosaccharides, disaccharides, nutritive/non-nutritive sugar alcohols, glycolytic and gluconeogenic metabolic intermediates for their cytoprotection against MPP+ in murine brain neuroblastoma cells. Several monosaccharides were effective against MMP+ (500 microM) including glucose, fructose and mannose, which restored cell viability to 109 +/- 5%, 70 +/- 5%, 99 +/- 3% of live controls, respectively. Slight protective effects were observed in the presence of 3-phosphoglyceric acid and glucose-6-phosphate; however, no protective effects were exhibited by galactose, sucrose, sorbitol, mannitol, glycerol or various gluconeogenic and ketogenic amino acids. On the other hand, fructose 1,6 bisphosphate and gluconeogenic energy intermediates [pyruvic acid, malic acid and phospho(enol)pyruvate (PEP)] were neuroprotective against MPP+. The gluconeogenic intermediates elevated intracellular levels of ATP and reduced propidium iodide (PI) nucleic acid staining to live controls, but did not alter the MPP(+)-induced loss of mitochondrial O2 consumption. These data indicate that malic acid, pyruvic acid and PEP contribute to anaerobic substrate level phosphorylation. The use of hydrazine sulfate to impede gluconeogenesis through PEP carboxykinase (PEPCK) inhibition heightened the protective effects of energy substrates possibly due to attenuated ATP demands from pyruvate carboxylase (PC) activity and pyruvate mitochondrial transport. It was concluded from these studies that several metabolic intermediates are effective in fueling anaerobic glycolysis during mitochondrial inhibition by MPP+.

1-Methyl-4-phenylpyridinium↗