Whole Body De Novo Amino Acid Synthesis in Type I (Insulin-dependent) Diabetes Studied With Stable Isotope-labeled Leucine, Alanine, and Glycine

We have investigated whether there is a net contribution of lysine synthesized de novo by the gastrointestinal microflora to lysine homeostasis in six adults. On two separate occasions an adequate diet was given for a total of 11 days, and a 24-h (12-h fast, 12-h fed) tracer protocol was performed on the last day, in which lysine turnover, oxidation, and splanchnic uptake were measured on the basis of intravenous and oral administration ofl-[1-13C]lysine andl-[6,6-2H2]lysine, respectively. [15N2]urea or15NH4Cl was ingested daily over the last 6 days to label microbial protein. In addition, seven ileostomates were studied with15NH4Cl. [15N]lysine enrichment in fecal and ileal microbial protein, as precursor for microbial lysine absorption, and in plasma free lysine was measured by gas chromatography-combustion-isotope ratio mass spectrometry. Differences in plasma [13C]- and [2H2]lysine enrichments during the 12-h fed period were observed between the two15N tracer studies, although the reason is unclear, and possibly unrelated to the tracer form per se. In the normal adults, after15NH4Cl and [15N2]urea intake, respectively, lysine derived from fecal microbial protein accounted for 5 and 9% of the appearance rate of plasma lysine. With ileal microbial lysine enrichment, the contribution of microbial lysine to plasma lysine appearance was 44%. This amounts to a gross microbial lysine contribution to whole body plasma lysine turnover of between 11 and 130 mg ⋅ kg−1 ⋅ day−1, depending on the [15N]lysine precursor used. However, insofar as microbial amino acid synthesis is accompanied by microbial breakdown of endogenous amino acids or their oxidation by intestinal tissues, this may not reflect a net increase in lysine absorption. Thus we cannot reliably estimate the quantitative contribution of microbial lysine to host lysine homeostasis with the present paradigm. However, the results confirm the significant presence of lysine of microbial origin in the plasma free lysine pool.

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Proteasome Inhibition in Brassica napus Roots Increases Amino Acid Synthesis to Offset Reduced Proteolysis

Plant and Cell Physiology ◽

10.1093/pcp/pcaa047 ◽

2020 ◽

Vol 61 (6) ◽

pp. 1028-1040

Author(s):

Dan Pereksta ◽

Dillon King ◽

Fahmida Saki ◽

Amith Maroli ◽

Elizabeth Leonard ◽

...

Keyword(s):

Amino Acid ◽

Brassica Napus ◽

De Novo ◽

Metabolic Response ◽

Sugar Metabolism ◽

Proteasome Inhibition ◽

Acid Synthesis ◽

Amino Acid Synthesis ◽

Amino Acid Depletion ◽

Metabolic Activities

Abstract Cellular homeostasis is maintained by the proteasomal degradation of regulatory and misfolded proteins, which sustains the amino acid pool. Although proteasomes alleviate stress by removing damaged proteins, mounting evidence indicates that severe stress caused by salt, metal(oids), and some pathogens can impair the proteasome. However, the consequences of proteasome inhibition in plants are not well understood and even less is known about how its malfunctioning alters metabolic activities. Lethality causes by proteasome inhibition in non-photosynthetic organisms stem from amino acid depletion, and we hypothesized that plants respond to proteasome inhibition by increasing amino acid biosynthesis. To address these questions, the short-term effects of proteasome inhibition were monitored for 3, 8 and 48 h in the roots of Brassica napus treated with the proteasome inhibitor MG132. Proteasome inhibition did not affect the pool of free amino acids after 48 h, which was attributed to elevated de novo amino acid synthesis; these observations coincided with increased levels of sulfite reductase and nitrate reductase activities at earlier time points. However, elevated amino acid synthesis failed to fully restore protein synthesis. In addition, transcriptome analysis points to perturbed abscisic acid signaling and decreased sugar metabolism after 8 h of proteasome inhibition. Proteasome inhibition increased the levels of alternative oxidase but decreased aconitase activity, most sugars and tricarboxylic acid metabolites in root tissue after 48 h. These metabolic responses occurred before we observed an accumulation of reactive oxygen species. We discuss how the metabolic response to proteasome inhibition and abiotic stress partially overlap in plants.

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Target of Rapamycin Inhibition in Chlamydomonas reinhardtii Triggers de Novo Amino Acid Synthesis by Enhancing Nitrogen Assimilation

The Plant Cell ◽

10.1105/tpc.18.00159 ◽

2018 ◽

Vol 30 (10) ◽

pp. 2240.1-2254 ◽

Cited By ~ 19

Author(s):

Umarah Mubeen ◽

Jessica Jüppner ◽

Jessica Alpers ◽

Dirk K. Hincha ◽

Patrick Giavalisco

Keyword(s):

Amino Acid ◽

Chlamydomonas Reinhardtii ◽

Nitrogen Assimilation ◽

De Novo ◽

Acid Synthesis ◽

Target Of Rapamycin ◽

Amino Acid Synthesis

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Mechanism of De Novo Branched-Chain Amino Acid Synthesis as an Alternative Electron Sink in Hypoxic Aspergillus nidulans Cells

Applied and Environmental Microbiology ◽

10.1128/aem.02135-09 ◽

2010 ◽

Vol 76 (5) ◽

pp. 1507-1515 ◽

Cited By ~ 36

Author(s):

Motoyuki Shimizu ◽

Tatsuya Fujii ◽

Shunsuke Masuo ◽

Naoki Takaya

Keyword(s):

Amino Acids ◽

Amino Acid ◽

Aspergillus Nidulans ◽

De Novo ◽

Acid Synthesis ◽

Branched Chain Amino Acids ◽

Amino Acid Synthesis ◽

Lactate Dehydrogenase Activity ◽

Branched Chain Amino Acid ◽

Branched Chain

ABSTRACT Although branched-chain amino acids are synthesized as building blocks of proteins, we found that the fungus Aspergillus nidulans excretes them into the culture medium under hypoxia. The transcription of predicted genes for synthesizing branched-chain amino acids was upregulated by hypoxia. A knockout strain of the gene encoding the large subunit of acetohydroxy acid synthase (AHAS), which catalyzes the initial reaction of the synthesis, required branched-chain amino acids for growth and excreted very little of them. Pyruvate, a substrate for AHAS, increased the amount of hypoxic excretion in the wild-type strain. These results indicated that the fungus responds to hypoxia by synthesizing branched-chain amino acids via a de novo mechanism. We also found that the small subunit of AHAS regulated hypoxic branched-chain amino acid production as well as cellular AHAS activity. The AHAS knockout resulted in higher ratios of NADH/NAD+ and NADPH/NADP+ under hypoxia, indicating that the branched-chain amino acid synthesis contributed to NAD+ and NADP+ regeneration. The production of branched-chain amino acids and the hypoxic induction of involved genes were partly repressed in the presence of glucose, where cells produced ethanol and lactate and increased levels of lactate dehydrogenase activity. These indicated that hypoxic branched-chain amino acid synthesis is a unique alternative mechanism that functions in the absence of glucose-to-ethanol/lactate fermentation and oxygen respiration.

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Glutamine nitrogen kinetics in insulin-dependent diabetic humans

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.1991.261.6.e713 ◽

1991 ◽

Vol 261 (6) ◽

pp. E713-E718 ◽

Cited By ~ 2

Author(s):

D. Darmaun ◽

M. Rongier ◽

J. Koziet ◽

J. J. Robert

Keyword(s):

Insulin Infusion ◽

De Novo ◽

Significant Rise ◽

Whole Body ◽

Type I ◽

Insulin Deficiency ◽

Appearance Rate ◽

Insulin Dependent ◽

Insulin Dependent Diabetic ◽

Infusion System

To assess the effect of insulin deficiency on whole body glutamine kinetics, five young adults with type I (insulin-dependent) diabetes received 4-h primed continuous infusions of L-[1-13C]leucine and L-[2-15N]glutamine in the postabsorptive state after blood glucose had been clamped overnight at either a normoglycemic level (approximately 85 mg/dl) or a moderate hyperglycemic level (approximately 260 mg/dl) by means of an automated glucose control insulin infusion system. The hyperglycemic state was associated with a significant rise in leucine level [from 165 +/- 23 to 242 +/- 62 (SD) microM], appearance rate (from 125 +/- 11 to 142 +/- 17 mumol.kg-1.h-1), and oxidation (from 27 +/- 10 to 31 +/- 10 mumol.kg-1.h-1). In contrast, neither the plasma level nor the appearance rate of glutamine (333 +/- 51 vs. 318 +/- 58 mumol.kg-1.h-1) was affected. We conclude that insulin deficiency resulting in moderate hyperglycemia induces a 13% rise in whole body proteolysis and yet does not stimulate glutamine de novo synthesis, despite increased precursor availability.

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A new stable isotope tracer technique to assess human neonatal amino acid synthesis

Journal of Pediatric Surgery ◽

10.1016/0022-3468(95)90496-4 ◽

1995 ◽

Vol 30 (9) ◽

pp. 1325-1329 ◽

Cited By ~ 16

Author(s):

Ronna G Miller ◽

Farook Jahoor ◽

Peter J Reeds ◽

William C Heird ◽

Tom Jaksic

Keyword(s):

Amino Acid ◽

Stable Isotope ◽

Acid Synthesis ◽

Tracer Technique ◽

Amino Acid Synthesis ◽

Stable Isotope Tracer ◽

Isotope Tracer

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Transient increase of de novo amino acid synthesis and its physiological significance in water-stressed white clover

Functional Plant Biology ◽

10.1071/fp05022 ◽

2005 ◽

Vol 32 (9) ◽

pp. 831 ◽

Cited By ~ 12

Author(s):

Bok-Rye Lee ◽

Woo-Jin Jung ◽

Kil-Yong Kim ◽

Jean-Christophe Avice ◽

Alain Ourry ◽

...

Keyword(s):

Protein Synthesis ◽

Amino Acid ◽

Water Deficit ◽

White Clover ◽

De Novo ◽

Acid Synthesis ◽

Ammonia Concentration ◽

De Novo Synthesis ◽

Amino Acid Synthesis ◽

De Novo Protein Synthesis

In white clover (Trifolium repens L. cv. Regal) the kinetics of de novo synthesis of amino acid and protein were compared by tracing 15N under well-watered (control) or water-deficit conditions. The physiological relationship between ammonia concentration, in response to the change in leaf water parameters, and de novo synthesis of amino acid and protein was also assessed. Leaf and root dry mass were not significantly affected for the first 3 d, whereas metabolic parameters such as total N and ammonia were significantly affected within the first day of water-deficit treatment. Inhibitory effect of water deficit on N acquisition from the soil was significant throughout the experimental period. Water deficit induced a significant increase in ammonia concentration in leaves during the first 3 d, and in roots for only the first day. In both leaves and roots, an increase in de novo amino acid synthesis, which peaked in leaves within the first 3 d of water-deficit treatment (Ψw ≥ –1.18 MPa), was observed. The rate of decrease in de novo protein synthesis gradually accelerated as the duration of the water-deficit treatment increased. There was a significant positive relationship between ammonia production and the increase in de novo amino acid synthesis during the first 3-d period, but not during the later period (day 3–day 7). This experiment clearly indicates that the increase in de novo amino acid synthesis caused by water deficit is a transient adaptive response occurring during the first few days and that it is associated with the increased ammonia concentrations, which in turn arise in response to a decrease in de novo protein synthesis.

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