Chemotaxis toward amino acids in Escherichia coli

Escherichia coli cells are shown to be attracted to the l-amino acids alanine, asparagine, aspartate, cysteine, glutamate, glycine, methionine, serine, and threonine, but not to arginine, cystine, glutamine, histidine, isoleucine, leucine, lysine, phenylalanine, tryptophan, tyrosine, or valine. Bacteria grown in a proline-containing medium were, in addition, attracted to proline. Chemotaxis toward amino acids is shown to be mediated by at least two detection systems, the aspartate and serine chemoreceptors. The aspartate chemoreceptor was nonfunctional in the aspartate taxis mutant, which showed virtually no chemotaxis toward aspartate, glutamate, or methionine, and reduced taxis toward alanine, asparagine, cysteine, glycine, and serine. The serine chemoreceptor was nonfunctional in the serine taxis mutant, which was defective in taxis toward alanine, asparagine, cysteine, glycine, and serine, and which showed no chemotaxis toward threonine. Additional data concerning the specificities of the amino acid chemoreceptors with regard to amino acid analogues are also presented. Finally, two essentially nonoxidizable amino acid analogues, alpha-aminoisobutyrate and alpha-methylaspartate, are shown to be attractants for E. coli, demonstrating that extensive metabolism of attractants is not required for amino acid taxis.     

Metabolism of amino acids in Ascaridia galli: decarboxylation reactions

Production of ¹⁴CO₂ from 12 carbon-labelled amino acids by Ascaridia galli was studied. Appreciable amounts of CO₂ were evolved from alanine, aspartate, glutamate, serine, leucine and valine by intestines, ovaries, cuticle and intact worms, in that order, but not from lysine, proline and tyrosine. Maximum CO₂ produced by whole worms was from serine, while with isolated organs it was from alanine. For cuticle, the decarboxylations of alanine, aspartate and glutamate were found to be associated with the mitochondrial fraction.     

Thermophilic anaerobic amino acid degradation: deamination rates and end-product formation

Anaerobic thermophilic degradation of several amino acids was studied in batch cultures using an inoculum from a steady-state semicontinuous enrichment culture. Experiments were done in the presence and absence of methanogenesis and known electron acceptors in the Stickland reaction. Methanogenesis was found to be crucial for the degradation of amino acids known to be oxidatively deaminated (leucine, valine and alanine). Other amino acids (serine, threonine, cysteine and methionine) were degraded under both methanogenic and non-methanogenic conditions. Degradation rates for these four amino acids were 1.3 to 2.2 times higher in cases where methanogenesis was active. The degradation rates of serine, threonine, cysteine and methionine were about twice as high as the rates of leucine, valine and alanine under methanogenic conditions. Inclusion of different electron acceptors, known to work in the Stickland reaction, did not enhance the degradation rates of any amino acid used nor did they alter the degradation patterns. Glycine was oxidatively deaminated to acetate, carbon dioxide, hydrogen and ammonium.     

Amino acid degradation by the mesophilic sulfate-reducing bacterium Desulfobacterium vacuolatum

Desulfobacterium vacuolatum strain IbRM was able to grow using casamino acids as a source of carbon, energy and nitrogen. Growth was accompanied by utilization of several amino acids and sulfide production. Proline and glutamate were used preferentially and to the greatest extent. Glycine, serine and alanine were used more slowly and only after proline and glutamate were used. Isoleucine, valine, leucine and aspartate decrease was slowest and occurred in a linear fashion throughout the growth phase. Amino acids used from casamino acids, excluding aspartate, were also used as single carbon, energy and nitrogen sources. As a single amino acid, aspartate could only be used as a nitrogen source. Aspartate was not used as an electron acceptor. No growth occurred on any amino acid in the absence of sulfate. As single substrates, isoleucine, proline and glutamate were oxidized without formation of acetate and with molar yields of 13.1, 9.4 and 7.7 g mol^–1, respectively. Received: 24 June 1997 / Accepted: 10 September 1997     

Nitrogen metabolism in tumor bearing mice

In experiments with whole animals infested with a highly malignant strain of Ehrlich ascites tumor cells, serial concentrations of amino acids were determined for host plasma, ascitic fluid, and tumor cells, throughout tumor development. Concentration gradients of glutamine, asparagine, valine, leucine, isoleucine, phenylalanine, tyrosine, histidine, tryptophan, arginine, serine, methionine, and taurine from the host plasma toward the ascitic liquid were established; while on the other hand, concentration gradients from the ascitic liquid toward the plasma were established for glutamate, aspartate, glycine, alanine, proline, and threonine. With the exception of aspartate the concentrations of these amino acids were highest inside the cells. Arginine was the only amino acid not detected in tumor cells. In vitro incubations of tumor cells in the presence of glutamine and/or glucose, as the energy and nitrogen sources, confirmed the amino acid fluxes previously deduced from the observed relative concentrations of amino acids in plasma, ascitic liquid, and tumor cells, suggesting that glutamate, alanine, aspartate, glycine, and serine can be produced by tumors. These findings support that changes in amino acid patterns occurring in the host system are related to tumor development.     

凤眼莲根分泌物氨基酸组分对根际肠杆菌属F2细菌的趋化作用

凤眼莲(Eichhornia crassipes)的根分泌物中含有Met等多种氨基酸,其中Met、GABA、Gly、Ala、Asp、Ser、Val和Leu(10^-7～10^-2mol·L^-1)均对凤眼莲的根际肠杆菌属F₂(Enterobacter sp.F₂)细菌有强烈的正趋化作用;Glu、Thr和His(10^-7～10^-3mol·L^-1)也对该菌有一定的正趋化作用;而Lys、Cys、Arg、Tyr、Pro、Asn、Gln、Ile、Phe和Typ则对该菌表现出一定的负趋化作用.对细菌的正趋化作用存在一个趋化物的最适浓度范围.具有正趋化作用的氨基酸在凤眼莲根际的浓度都较高,而具有负趋化作用的浓度则较低,这正是凤眼莲与该根际细菌结合为根际微生态系统的原因之一.     

Metabolism of amino acids in Ascaridia galli: transamination

Ascaridia galli, using 2-oxoglutarate as an acceptor, transaminates alanine and aspartate at significantly high rates. Among other amino acids valine, phenylalanine, leucine, isoleucine, arginine, tyrosine and methionine are metabolised at moderate rates while lysine, serine, threonine, cysteine, glycine, histidine, tryptophan, DOPA and GABA appear to be inert in this respect. Body parts mimic the whole worm in the pattern of transamination of various amino acids with the exception of methionine. Alanine, aspartate and glutamate may transfer their amino group also to pyruvate and oxaloacetate. Alanine and aspartate: 2-oxoglutarate transaminases are located mainly in the cytosol and mitochondrial fractions.     

Alteration in the Amino Acid Content of Yeast During Growth Under Various Nutritional Conditions

Yeast cells grown under optimal and suboptimal concentrations of biotin were analyzed for the amino acid content of their soluble pool and cellular protein. Optimally grown yeast cells exhibited a maximum amino acid content after 18 hr of growth. Biotin-deficient cells were depleted of all amino acids at 26 and 43 hr, with alanine, arginine, aspartate, cysteine, glutamate, isoleucine, leucine, lysine, methionine, serine, threonine, and valine being present in less than half the concentration observed in biotin-optimal cells. At early time intervals, the amino acid pool of biotin-deficient yeast contained lower concentrations of all amino acids except alanine. After more prolonged incubation, several amino acids accumulated in the pool of biotin-deficient yeast, but citrulline and ornithine accumulated to appreciable levels. The addition of aspartate to the growth medium resulted in a decrease in the amino acid content of biotin-optimal cells but caused a marked increase in the concentration of amino acids in biotin-deficient cells. The pools of biotin-deficient yeast grown in the presence of aspartate displayed a marked reduction in every amino acid with the exception of aspartate itself. These data provide evidence that the amino acid content of yeast cells and their free amino acid pools are markedly affected by biotin deficiency as well as by supplementation with aspartate, indicating that aspartate plays a major role in the nitrogen economy of yeast under both normal as well as abnormal nutritional conditions.     

Factors affecting protein synthesis during biotin deficiency inAspergillus nidulans

Biotin deficiency in Aspergillus nidulans resulted in a 70% increase of the protein content and increased levels of free and bound aspartate, glutamate, serine, leucine and methionine. Likewise, the activities of NADP+ glutamate dehydrogenase, NAD+ gluatmate dehydrogenase, aspartate aminotransferase and alanine aminotransferase were significantly increased. The total RNA content increased while the DNA content was unaffected. The rRNA/tRNA ratio remained higher in biotin-deficient cells. Supplementation of glutamate, aspartate, serine, leucine and methionine to the culture medium raised the rRNA/tRNA ratio, and the difference observed in the qualitative and the quantitative patterns of protein and dry cell mass between normal and biotin-deficient cultures was abolished.     

Changes in plasma amino acid concentrations with increasing age in patients with propionic acidemia

The objective of the study is to analyze plasma amino acid concentrations in propionic acidemia (PA) for the purpose of elucidating possible correlations between propionyl-CoA carboxylase deficiency and distinct amino acid behavior. Plasma concentrations of 19 amino acids were measured in 240 random samples from 11 patients (6 families) with enzymatically and/or genetically proven propionic acidemia (sampling period, January 2001–December 2007). They were compared with reference values from the literature and correlated with age using the Pearson correlation coefficient test. Decreased plasma concentrations were observed for glutamine, histidine, threonine, valine, isoleucine, leucine, phenylalanine and arginine. Levels of glycine, alanine and aspartate were elevated, while values of serine, asparagine, ornithine and glutamate were normal. For lysine, proline and methionine a clear association was not possible. Significant correlations with age were observed for 13 amino acids (positive correlation: asparagine, glutamine, proline, alanine, histidine, threonine, methionine, arginine; negative correlation: leucine, phenylalanine, ornithine, glutamate and aspartate). This study gives new insight over long-term changes in plasma amino acid concentrations and may provide options for future therapies (e.g., substitution of anaplerotic substances) in PA patients.     

The uptake of amino acids from a chemically defined medium byFusobacterium species

The uptake of amino acids from a chemically defined medium was determined for various species ofFusobacterium. Reference strains utilized a wider range and higher levels of amino acids than clinical isolates. Among the acidic and basic amino acids, arginine, histidine, and glutamate were used by most species. In general, nonpolar neutral amino acids such as alanine, valine, leucine, isoleucine, proline, and methionine were poorly utilized. Apart from glycine and tyrosine, the neutral but polar amino acids such as serine, cysteine, and asparagine were incorporated at significantly high levels. ThusFusobacterium species, unlike several Gram-negative anaerobic bacteria, have the capacity to utilize both free amino acids and peptides as energy sources and may partly account for the wide distribution of these species in areas of tissue degradation.     

Amino Acid Metabolism of Lemna minor L. : III. Responses to Aminooxyacetate

Aminooxyacetate, a known inhibitor of transaminase reactions and glycine decarboxylase, promotes rapid depletion of the free pools of serine and aspartate in nitrate grown Lemna minor L. This compound markedly inhibits the methionine sulfoximine-induced accumulation of free ammonium ions and greatly restricts the methionine sulfoximine-induced depletion of amino acids such as glutamate, alanine, and asparagine. These results suggest that glutamate, alanine, and asparagine are normally catabolized to ammonia by transaminase-dependent pathways rather than via dehydrogenase or amidohydrolase reactions. Aminooxyacetate does not inhibit the methionine sulfoximine-induced irreversible deactivation of glutamine synthetase in vivo, indicating that these effects cannot be simply ascribed to inhibition of methionine sulfoximine uptake by amino-oxyacetate. This transaminase inhibitor promotes extensive accumulation of several amino acids including valine, leucine, isoleucine, alanine, glycine, threonine, proline, phenylalanine, lysine, and tyrosine. Since the aminooxyacetate induced accumulations of valine, leucine, and isoleucine are not inhibited by the branched-chain amino acid biosynthesis inhibitor, chlorsulfuron, these amino acid accumulations most probably involve protein turnover. Depletions of soluble protein bound amino acids are shown to be approximately stoichiometric with the free amino acid pool accumulations induced by aminooxyacetate. Aminooxyacetate is demonstrated to inhibit the chlorsulfuron-induced accumulation of α-amino-n-butyrate in L. minor, supporting the notion that this amino acid is derived from transamination of 2-oxobutyrate.     

Amino Acid Metabolism of Lemna minor L. : I. Responses to Methionine Sulfoximine

When Lemna minor L. is supplied with the potent inhibitor of glutamine synthetase, methionine sulfoximine, rapid changes in free amino acid levels occur. Glutamine, glutamate, asparagine, aspartate, alanine, and serine levels decline concomitantly with ammonia accumulation. However, not all free amino acid pools deplete in response to this inhibitor. Several free amino acids including proline, valine, leucine, isoleucine, threonine, lysine, phenylalanine, tyrosine, histidine, and methionine exhibit severalfold accumulations within 24 hours of methionine sulfoximine treatment. To investigate whether these latter amino acid accumulations result from de novo synthesis via a methionine sulfoximine insensitive pathway of ammonia assimilation (e.g. glutamate dehydrogenase) or from protein turnover, fronds of Lemna minor were prelabeled with [¹⁵N]H₄⁺ prior to supplying the inhibitor. Analyses of the ¹⁵N abundance of free amino acids suggest that protein turnover is the major source of these methionine sulfoximine induced amino acid accumulations. Thus, the pools of valine, leucine, isoleucine, proline, and threonine accumulated in response to the inhibitor in the presence of [¹⁵N]H₄⁺, are ¹⁴N enriched and are not apparently derived from ¹⁵N-labeled precursors. To account for the selective accumulation of amino acids, such as valine, leucine, isoleucine, proline, and threonine, it is necessary to envisage that these free amino acids are relatively poorly catabolized in vivo. The amino acids which deplete in response to methionine sulfoximine (i.e. glutamate, glutamine, alanine, aspartate, asparagine, and serine) are all presumably rapidly catabolized to ammonia, either in the photorespiratory pathway or by alternative routes.     

Control of proteolysis in perifused rat hepatocytes

The mechanism by means of which amino acids inhibit intrahepatic protein degradation has been studied in perifused rat hepatocytes. Proteolysis was extremely sensitive to inhibition by low concentrations of amino acids. A mixture of 0.5 mM leucine and 1-2 mM alanine, concentrations found in the portal vein of the rat after feeding, inhibited proteolysis to the same extent as a complete physiological mixture of amino acids. Inhibition by these two amino acids was accompanied by a rise in the intracellular concentrations of glutamate and aspartate, and was largely prevented by addition of glucagon, by addition of the transaminase inhibitor aminooxyacetate, or by omission of K+. Acceleration of proteolysis by K+ depletion was accompanied by a fall in intracellular glutamate caused by an increased rate of transport of this amino acid to the extracellular fluid. It is concluded that intracellular leucine, glutamate and aspartate are important elements in the control of hepatic protein degradation.     

Metabolic footprinting of the anaerobic bacterium Fusobacterium varium using 1H NMR spectroscopy

Metabolic footprinting of the anaerobic bacterium Fusobacterium varium demonstrated the accumulation of six carboxylic acids as metabolic end-products and revealed specific growth requirements and utilization capabilities towards amino acids. Guided by (1)H NMR determinations of residual amino acids in spent medium, a modified chemically defined minimal medium (CDMM*) was developed by minimizing the amino acid composition while satisfying nutritional requirements to support abundant growth of F. varium. Quantitative determinations of carboxylate salts and residual substrates were readily performed by (1)H NMR analysis of lyophilized residues from CDMM* cultures without interference from initial medium components. Only small concentrations of alanine, arginine, glycine, isoleucine, leucine, methionine, proline and valine were required to support growth of F. varium, whereas larger quantities of aspartate, asparagine, cysteine, glutamine, glutamate, histidine, lysine, serine and threonine were utilized, most likely as energy sources. Both bacterial growth and the distribution of carboxylate end-products depended on the composition of the chemically defined medium. In cultures provided with glucose as the primary energy source, the accumulation of butyrate and lactate correlated with growth, consistent with the regeneration of reduced coenzyme formed by the oxidative steps of glucose catabolism.     

Use of 3H and 14C double-labeled glucose to assess in vivo pathways of amino acid biosynthesis in Escherichia coli.

3H and 14C tracing data concerning amino acid biosynthetic pathways in Escherichia coli K12 are presented. Thirteen acidic and neutral amino acids were isolated from protein hydrolysates of wild type E. coli K12 grown aerobically or anaerobically in the presence of [U-14C]glucose together with [1-3H]glucose, [3-3H]glucose, [4-3H]glucose, or [6-3H]glucose. The observed 3H/14C counts of the amino acids were compared with the ratios expected on the basis of the input substrate specific activities and present understanding of biosynthetic pathways. For nine amino acids, serine, valine, leucine, threonine, isoleucine, glycine, glutamate, proline, and phenylalanine, the agreement between anticipated and observed specific activities was satisfactory. For the remaining four, methionine, alanine, aspartate, and (in cells labeled with [3-3H]glucose) tyrosine, the anticipated and observed specific activities differed markedly. For alanine, aspartate, and tyrosine, the differences are probably due to exchange of tritium in the course of biosynthesis; for methionine, it may be that there is a principle source of the methyl group other than carbon 3 of serine.     

Glucose and amino acid metabolism in neonatal rat skeletal muscle in tissue culture

The characteristics of glucose and amino acid metabolism over a 98-hour incubation period were studied in a primary culture of neonatal rat skeletal muscle cells. The cells formed large myotubes in culture, were spontaneously highly contractile, and had cell phosphocreatine levels exceeding ATP concentrations. Medium glucose fell from 7.2±0.2 to 1.5±0.1 mM between 0 and 98 hours; intracellular glucose was readily detectable, indicating glycolysis was limited by phosphorylation, not glucose transport. Alanine levels in the medium increased from 0.06±0.01 to 0.82±0.04 mM between 0 and 48 hours and decreased to 0.72±0.04 mM by 98 hours. The period of net alanine production correlated with the rise in the cell mass action ratio of the alanine aminotransferase reaction. Cell aspartate, glutamate, and calculated oxalacetate levels were inversely related to the cell NADH/NAD⁺ ratio, as represented by the intracellular lactate/pyruvate ratio (r=0.78–0.88). The branched chain amino acids (leucine, isoleucine, valine) were actively utilized, e.g., medium leucine fell from 0.70±0.01 to 0.30±0.06 mM between 0 and 98 hours. In addition, arginine and serine consumption was observed in conjunction with ornithine, proline, and glycine production. Conclusions: (1) A major driving force for the high rates of alanine production by skeletal muscle cells in tissue culture is the active utilization of branched chain amino acids. (2) Intracellular aspartate and glutamate pools are linked, probably via the malate-aspartate shuttle, to the cell NADH/NAD⁺ redox state. (3) Muscle cells in tissue culture metabolize significant amounts of arginine and serine in association with the production of ornithine and proline, and these pathways may possibly be related to creatine production.     

l-Glutamine regulates amino acid utilization by intestinal bacteria

Catabolism of amino acids (AA) by intestinal bacteria greatly affects their bioavailability in the systemic circulation and the health of animals and humans. This study tests the novel hypothesis that l-glutamine regulates AA utilization by luminal bacteria of the small intestine. Pure bacterial strains (Streptococcus sp., Escherichia coli and Klebsiella sp.) and mixed bacterial cultures derived from the jejunum or ileum of pigs were cultured in the presence of 0–5 mM l-glutamine under anaerobic conditions. After 3 h of incubation, samples were taken for the determination of AA utilization. Results showed concentration-dependent increases in the utilization of glutamine in parallel with the increased conversion of glutamine into glutamate in all the bacteria. Complete utilization of asparagine, aspartate and serine was observed in pure bacterial strains after the 3-h incubation. The addition of glutamine reduced the net utilization of asparagine by both jejunal and ileal mixed bacteria. Net utilization of lysine, leucine, valine, ornithine and serine by jejunal or ileal mixed bacteria decreased with the addition of glutamine in a concentration-dependent manner. Collectively, glutamine dynamically modulates the bacterial metabolism of the arginine family of AA as well as the serine and aspartate families of AA and reduced the catabolism of most AA (including nutritionally essential and nonessential AA) in jejunal or ileal mixed bacteria. The beneficial effects of glutamine on gut nutrition and health may involve initiation of the signaling pathways related to AA metabolism in the luminal bacteria of the small intestine.     

Extracellular Levels of Amino Acids and Choline in Human High Grade Gliomas: An Intraoperative Microdialysis Study

The concentrations of endogenous amino acids and choline in the extracellular fluid of human cerebral gliomas have been measured, for the first time, by in vivo microdialysis. Glioblastoma growth was associated with increased concentrations of choline, GABA, isoleucine, leucine, lysine, phenylalanine, taurine, tyrosine, and valine. There was no difference between grade III and grade IV tumors in the concentrations of phenylalanine, isoleucine, tyrosine, valine, and lysine, whereas the concentrations of choline, aspartate, taurine, GABA, leucine, and glutamate were significantly different in the two tumor-grade subgroups. In contrast to the other compounds, the concentration of glutamate was decreased in glioma. The parenchyma adjacent to the tumor showed significant changes only in the extracellular concentration of glutamate, isoleucine, and valine. The concentrations of choline and the amino acids, glutamate, leucine, taurine, and tyrosine showed significant positive correlations with the degree of cell proliferation. Epilepsy, which is relatively common in subjects with gliomas, was shown to be a significant confounding variable when the extracellular concentrations of aspartate, glutamate and GABA were considered.     

The effects of starvation on plasma free amino acid and glucose concentrations in lake sturgeon

The effect of starvation on the metabolism of the lake sturgeon Acipenser fulvescens was examined by measuring haematocrit, plasma glucose concentrations, and plasma free amino acids. Plasma was sampled on day 0, 10, 20, 45 and 60 of a 60-day starvation period. Haematocrit was observed to decrease with starvation indicating a decreased oxygen carrying capacity of the blood. Plasma glucose levels differed only at day 10, with a decrease in blood glucose level in the starved group. No differences were detected between groups for alanine, aspartate, and serine, while elevated levels were observed for glutamine throughout the experiment. An increase in arginine, tyrosine, valine, methionine, tryptophan, phenylalanine, glutamate, glycine, isoleucine, histidine and leucine, concentrations were observed after 45 days of starvation. The maintenance, or increased plasma levels, of glucogenic amino acids in combination with the maintenance of blood glucose concentrations indicates active gluconeogenic processes in the liver supported by muscle proteolysis.