scholarly journals Metabolic flux from the Krebs cycle to glutamate transmission tunes a neural brake on seizure onset

PLoS Genetics ◽  
2021 ◽  
Vol 17 (10) ◽  
pp. e1009871
Author(s):  
Jiwon Jeong ◽  
Jongbin Lee ◽  
Ji-hyung Kim ◽  
Chunghun Lim
Keyword(s):  
Metabolic Flux ◽  
Krebs Cycle ◽  
Seizure Onset ◽  
Glutamatergic Neurons ◽  
Citrate Transporter ◽  
Neurodevelopmental Deficits ◽  
Glutamate Transmission ◽  
Underlying Mechanisms ◽  
Glutamate Biosynthesis ◽  
Rate Limiting

Kohlschütter-Tönz syndrome (KTS) manifests as neurological dysfunctions, including early-onset seizures. Mutations in the citrate transporter SLC13A5 are associated with KTS, yet their underlying mechanisms remain elusive. Here, we report that a Drosophila SLC13A5 homolog, I’m not dead yet (Indy), constitutes a neurometabolic pathway that suppresses seizure. Loss of Indy function in glutamatergic neurons caused “bang-induced” seizure-like behaviors. In fact, glutamate biosynthesis from the citric acid cycle was limiting in Indy mutants for seizure-suppressing glutamate transmission. Oral administration of the rate-limiting α-ketoglutarate in the metabolic pathway rescued low glutamate levels in Indy mutants and ameliorated their seizure-like behaviors. This metabolic control of the seizure susceptibility was mapped to a pair of glutamatergic neurons, reversible by optogenetic controls of their activity, and further relayed onto fan-shaped body neurons via the ionotropic glutamate receptors. Accordingly, our findings reveal a micro-circuit that links neural metabolism to seizure, providing important clues to KTS-associated neurodevelopmental deficits.

scholarly journals Metabolome and proteome analyses reveal transcriptional misregulation in glycolysis of engineered E. coli

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Chun-Ying Wang ◽  
Martin Lempp ◽  
Niklas Farke ◽  
Stefano Donati ◽  
Timo Glatter ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Binding Site ◽  
Metabolic Pathways ◽  
Glycerol Production ◽  
Inducible Promoters ◽  
E Coli ◽  
Inhibition Of Glycolysis ◽  
Underlying Mechanisms ◽  
Rate Limiting ◽  
Engineered Bacteria

AbstractSynthetic metabolic pathways are a burden for engineered bacteria, but the underlying mechanisms often remain elusive. Here we show that the misregulated activity of the transcription factor Cra is responsible for the growth burden of glycerol overproducing E. coli. Glycerol production decreases the concentration of fructose-1,6-bisphoshate (FBP), which then activates Cra resulting in the downregulation of glycolytic enzymes and upregulation of gluconeogenesis enzymes. Because cells grow on glucose, the improper activation of gluconeogenesis and the concomitant inhibition of glycolysis likely impairs growth at higher induction of the glycerol pathway. We solve this misregulation by engineering a Cra-binding site in the promoter controlling the expression of the rate limiting enzyme of the glycerol pathway to maintain FBP levels sufficiently high. We show the broad applicability of this approach by engineering Cra-dependent regulation into a set of constitutive and inducible promoters, and use one of them to overproduce carotenoids in E. coli.

Metabolism of totally ischemic excised dog heart I. Construction of a computer model

1979 ◽  
Vol 237 (5) ◽  
pp. R318-R326 ◽  
Author(s):  
Murray J. Achs ◽  
David Garfinkel
Keyword(s):  
Computer Model ◽  
Metabolic Regulation ◽  
Krebs Cycle ◽  
The Body ◽  
Heavy Sedation ◽  
Dog Heart ◽  
Rat Hearts ◽  
Heart Preparation ◽  
Time Courses ◽  
Underlying Mechanisms

Construction and fit to the experimental data of a computer model of glycolysis, the Krebs cycle, and related metabolism in an ischemic dog heart preparation, involving 122 metabolites, 65 enzymes, and 406 chemical reactions, is described. The experimental preparation simulated is a dog heart excised from the body, placed in a beaker of Tyrode's solution, and sampled for 100 min; the model required only moderate modification from models representing perfused rat hearts, and little modification from a model of another ischemic dog heart preparation. Common underlying mechanisms for the ischemia are indicated, although this preparation appears to evolve more slowly with time, perhaps owing to heavy sedation and diffusion-limited transport. Lactate is, at first, exported and then accumulates intracellularly; pH falls, but not as much in the mitochondria as the cytoplasm; redox couples go reduced, but with counterintuitive time courses; calcium phosphate is calculated to precipitate, as often observed in cardiac ischemia enzymes; glycolysis; simulation; metabolic regulation Submitted on October 24, 1978 Accepted on May 21, 1979

scholarly journals Advances in the Metabolic Engineering of Escherichia coli for the Manufacture of Monoterpenes

Catalysts ◽  
2019 ◽  
Vol 9 (5) ◽  
pp. 433 ◽  
Author(s):  
Si-si Xie ◽  
Lingyun Zhu ◽  
Xin-yuan Qiu ◽  
Chu-shu Zhu ◽  
Lv-yun Zhu
Keyword(s):  
Large Scale ◽  
Metabolic Flux ◽  
Process Development ◽  
Pathway Engineering ◽  
Scale Production ◽  
Future Studies ◽  
E Coli ◽  
Large Scale Production ◽  
Rate Limiting ◽  
Dependent Pathway

Monoterpenes are commonly applied as pharmaceuticals and valuable chemicals in various areas. The bioproduction of valuable monoterpenes in prokaryotic microbial hosts, such as E. coli, has progressed considerably thanks to the development of different outstanding approaches. However, the large-scale production of monoterpenes still presents considerable limitations. Thus, process development warrants further investigations. This review discusses the endogenous methylerythritol-4-phosphate-dependent pathway engineering and the exogenous mevalonate-dependent isoprenoid pathway introduction, as well as the accompanied optimization of rate-limiting enzymes, metabolic flux, and product toxicity tolerance. We suggest further studies to focus on the development of systematical, integrational, and synthetic biological strategies in light of the inter disciplines at the cutting edge. Our review provides insights into the current advances of monoterpene bioengineering and serves as a reference for future studies to promote the industrial production of valuable monoterpenes.

scholarly journals In Vivo Detection of Brain Krebs Cycle Intermediate by Hyperpolarized Magnetic Resonance

2012 ◽  
Vol 32 (12) ◽  
pp. 2108-2113 ◽  
Author(s):  
Mor Mishkovsky ◽  
Arnaud Comment ◽  
Rolf Gruetter
Keyword(s):  
Magnetic Resonance ◽  
Krebs Cycle ◽  
Tca Cycle ◽  
Energy Regulation ◽  
Biochemical Processes ◽  
In Vivo Detection ◽  
Intact Brain ◽  
Tca Cycle Intermediates ◽  
Rate Limiting

The Krebs (or tricarboxylic acid (TCA)) cycle has a central role in the regulation of brain energy regulation and metabolism, yet brain TCA cycle intermediates have never been directly detected in vivo. This study reports the first direct in vivo observation of a TCA cycle intermediate in intact brain, namely, 2-oxoglutarate, a key biomolecule connecting metabolism to neuronal activity. Our observation reveals important information about in vivo biochemical processes hitherto considered undetectable. In particular, it provides direct evidence that transport across the inner mitochondria membrane is rate limiting in the brain. The hyperpolarized magnetic resonance protocol designed for this study opens the way to direct and real-time studies of TCA cycle kinetics.

scholarly journals Transcriptional and Metabolic Responses of Bacillus subtilis to the Availability of Organic Acids: Transcription Regulation Is Important but Not Sufficient To Account for Metabolic Adaptation

2006 ◽  
Vol 73 (2) ◽  
pp. 499-507 ◽  
Author(s):  
Oliver Schilling ◽  
Oliver Frick ◽  
Christina Herzberg ◽  
Armin Ehrenreich ◽  
Elmar Heinzle ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Organic Acids ◽  
Pentose Phosphate Pathway ◽  
Metabolic Regulation ◽  
Metabolic Flux ◽  
Krebs Cycle ◽  
Pentose Phosphate ◽  
Metabolic Adaptation ◽  
Acetyl Coa ◽  
Additional Level

ABSTRACT The soil bacterium Bacillus subtilis can use sugars or organic acids as sources of carbon and energy. These nutrients are metabolized by glycolysis, the pentose phosphate pathway, and the Krebs citric acid cycle. While the response of B. subtilis to the availability of sugars is well understood, much less is known about the changes in metabolism if organic acids feeding into the Krebs cycle are provided. If B. subtilis is supplied with succinate and glutamate in addition to glucose, the cells readjust their metabolism as determined by transcriptome and metabolic flux analyses. The portion of glucose-6-phosphate that feeds into the pentose phosphate pathway is significantly increased in the presence of organic acids. Similarly, important changes were detected at the level of pyruvate and acetyl coenzyme A (acetyl-CoA). In the presence of organic acids, oxaloacetate formation is strongly reduced, whereas the formation of lactate is significantly increased. The alsSD operon required for acetoin formation is strongly induced in the presence of organic acids; however, no acetoin formation was observed. The recently discovered phosphorylation of acetolactate decarboxylase may provide an additional level of control of metabolism. In the presence of organic acids, both types of analyses suggest that acetyl-CoA was catabolized to acetate rather than used for feeding the Krebs cycle. Our results suggest that future work has to concentrate on the posttranslational mechanisms of metabolic regulation.

scholarly journals HMGCR inhibition stabilizes the glycolytic enzyme PKM2 to support the growth of renal cell carcinoma

PLoS Biology ◽  
2021 ◽  
Vol 19 (4) ◽  
pp. e3001197
Author(s):  
Jiajun Huang ◽  
Xiaoyu Zhao ◽  
Xiang Li ◽  
Jiwei Peng ◽  
Weihao Yang ◽  
...  
Keyword(s):  
Renal Cell Carcinoma ◽  
Cell Carcinoma ◽  
Renal Cell ◽  
Tumor Development ◽  
Serum Levels ◽  
Glycolytic Enzyme ◽  
Hsp90 Expression ◽  
Cholesterol Biosynthetic Pathway ◽  
Underlying Mechanisms ◽  
Rate Limiting

Renal cell carcinoma (RCC) is responsible for most cases of the kidney cancer. Previous research showed that low serum levels of cholesterol level positively correlate with poorer RCC-specific survival outcomes. However, the underlying mechanisms and functional significance of the role of cholesterol in the development of RCC remain obscure. 3-Hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR) plays a pivotal role in RCC development as it is the key rate-limiting enzyme of the cholesterol biosynthetic pathway. In this study, we demonstrated that the inhibition of HMGCR could accelerate the development of RCC tumors by lactate accumulation and angiogenesis in animal models. We identified that the inhibition of HMGCR led to an increase in glycolysis via the regulated HSP90 expression levels, thus maintaining the levels of a glycolysis rate-limiting enzyme, pyruvate kinase M2 (PKM2). Based on these findings, we reversed the HMGCR inhibition-induced tumor growth acceleration in RCC xenograft mice by suppressing glycolysis. Furthermore, the coadministration of Shikonin, a potent PKM2 inhibitor, reverted the tumor development induced by the HMGCR signaling pathway.

scholarly journals Overproduction of Trehalose: Heterologous Expression of Escherichia coli Trehalose-6-Phosphate Synthase and Trehalose-6-Phosphate Phosphatase in Corynebacterium glutamicum

2004 ◽  
Vol 70 (1) ◽  
pp. 370-376 ◽  
Author(s):  
Leandro Padilla ◽  
Reinhard Krämer ◽  
Gregory Stephanopoulos ◽  
Eduardo Agosin
Keyword(s):  
Escherichia Coli ◽  
Heterologous Expression ◽  
Corynebacterium Glutamicum ◽  
Metabolic Flux ◽  
Krebs Cycle ◽  
Exponential Growth Phase ◽  
Fourfold Increase ◽  
Potential Applications ◽  
Defined Culture ◽  
Phosphate Synthase

ABSTRACT Trehalose is a disaccharide with potential applications in the biotechnology and food industries. We propose a method for industrial production of trehalose, based on improved strains of Corynebacterium glutamicum. This paper describes the heterologous expression of Escherichia coli trehalose-synthesizing enzymes trehalose-6-phosphate synthase (OtsA) and trehalose-6-phosphate phosphatase (OtsB) in C. glutamicum, as well as its impact on the trehalose biosynthetic rate and metabolic-flux distributions, during growth in a defined culture medium. The new recombinant strain showed a five- to sixfold increase in the activity of OtsAB pathway enzymes, compared to a control strain, as well as an almost fourfold increase in the trehalose excretion rate during the exponential growth phase and a twofold increase in the final titer of trehalose. The heterologous expression described resulted in a reduced specific glucose uptake rate and Krebs cycle flux, as well as reduced pentose pathway flux, a consequence of downregulated glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase. The results proved the suitability of using the heterologous expression of Ots proteins in C. glutamicum to increase the trehalose biosynthetic rate and yield and suggest critical points for further improvement of trehalose overproduction in C. glutamicum.

Mechanism of the inhibition of myocardial protein synthesis during oxygen deprivation

1976 ◽  
Vol 230 (1) ◽  
pp. 120-126 ◽  
Author(s):  
M Lesch ◽  
H Taegtmeyer ◽  
MB Peterson ◽  
R Vernick
Keyword(s):  
Protein Synthesis ◽  
Metabolic Regulation ◽  
Metabolic Flux ◽  
Adenine Nucleotide ◽  
Krebs Cycle ◽  
High Energy ◽  
Metabolic Inhibitors ◽  
High Energy Phosphate ◽  
Inhibition Of Protein Synthesis ◽  
Phosphate Depletion

Inhibition of protein synthesis during anoxia in the isolated rabbit right ventricular papillary muscle preparation is totally reversible for up to 2 h if glucose concentration is increased during anoxia. The degree of inhibition of protein synthesis during anoxia is, however, not altered by the presence of increased glucose. Thus inhibition of myocardial protein synthesis induced by anoxia need not be related to irreversible disruption of cellular integrity but may represent metabolic regulation of the synthesis. Tissue content of ATP, ADP, AMP, CP, and lactate and phenylalanine incorporation into protein were measured in individual papillary muscles incubated with varying degrees of O2 deprivation and varying substrates and metabolic inhibitors to determine if the inhibition during anoxia could be ascribed to alterations in tissue high-energy phosphate, adenine nucleotide levels, or rate of metabolic flux through the glycolytic and/or Krebs cycle. Protein synthesis was inhibited in muscles incubated in 15 mM glucose during anoxia despite the fact that in the presence of increased glucose, tissue levels of ATP, ADP, and AMP were equal to that of controls. Protein synthesis was normal in muscles made sufficiently hypoxic so that ATP and CP were significantly decreased and lactate increased. Inhibition of Krebs cycle activity with pentenoate failed to effect the rate of protein synthesis. We conclude that anoxic inhibition of myocardial protein synthesis is due neither to high-energy phosphate depletion nor inhibition of Krebs cycle acitivity. The possibility remains that the inhibition may be related to accumulation of glycolytic intermediates or by-products other than lactate.

scholarly journals Overproduction of Geranylgeraniol by Metabolically Engineered Saccharomyces cerevisiae

2009 ◽  
Vol 75 (17) ◽  
pp. 5536-5543 ◽  
Author(s):  
Kenro Tokuhiro ◽  
Masayoshi Muramatsu ◽  
Chikara Ohto ◽  
Toshiya Kawaguchi ◽  
Shusei Obata ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Metabolic Flux ◽  
Mevalonate Pathway ◽  
Fusion Gene ◽  
Pharmaceutical Products ◽  
Test Tube ◽  
Ethanol Solution ◽  
Multicopy Integration ◽  
Engineered Saccharomyces Cerevisiae ◽  
Rate Limiting

ABSTRACT (E, E, E)-Geranylgeraniol (GGOH) is a valuable starting material for perfumes and pharmaceutical products. In the yeast Saccharomyces cerevisiae, GGOH is synthesized from the end products of the mevalonate pathway through the sequential reactions of farnesyl diphosphate synthetase (encoded by the ERG20 gene), geranylgeranyl diphosphate synthase (the BTS1 gene), and some endogenous phosphatases. We demonstrated that overexpression of the diacylglycerol diphosphate phosphatase (DPP1) gene could promote GGOH production. We also found that overexpression of a BTS1-DPP1 fusion gene was more efficient for producing GGOH than coexpression of these genes separately. Overexpression of the hydroxymethylglutaryl-coenzyme A reductase (HMG1) gene, which encodes the major rate-limiting enzyme of the mevalonate pathway, resulted in overproduction of squalene (191.9 mg liter−1) rather than GGOH (0.2 mg liter−1) in test tube cultures. Coexpression of the BTS1-DPP1 fusion gene along with the HMG1 gene partially redirected the metabolic flux from squalene to GGOH. Additional expression of a BTS1-ERG20 fusion gene resulted in an almost complete shift of the flux to GGOH production (228.8 mg liter−1 GGOH and 6.5 mg liter−1 squalene). Finally, we constructed a diploid prototrophic strain coexpressing the HMG1, BTS1-DPP1, and BTS1-ERG20 genes from multicopy integration vectors. This strain attained 3.31 g liter−1 GGOH production in a 10-liter jar fermentor with gradual feeding of a mixed glucose and ethanol solution. The use of bifunctional fusion genes such as the BTS1-DPP1 and ERG20-BTS1 genes that code sequential enzymes in the metabolic pathway was an effective method for metabolic engineering.

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