A glutamate dehydrogenase-based method for the assay ofl-glutamic acid: Formation of pyridine nucleotide fluorescent derivatives

Mora, Miguel A.; Méndez-Franco, J; Salceda, Rocı́o; Riesgo-Escovar, Juan R.

doi:10.1016/0003-2697(89)90425-9

Cited by 20 publications

(9 citation statements)

References 13 publications

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“…In the absence of a mass spectrometer, biochemical assays could be used to quantify glutamate and glutamine. An assay for glutamine that relies on hydrolysis of glutamine to glutamate and quantification of glutamate as well as an assay for glutamate that relies on oxidation of glutamate to alpha-ketoglutarate and quantification of the formed NADH 35 are commercially available.…”

Section: Discussionmentioning

confidence: 99%

Glutamate enrichment as new diagnostic opportunity in breast cancer

Budczies

Pfitzner

Győrffy

et al. 2014

Intl Journal of Cancer

110

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Exogenous glutamine is an important source of energy and molecular building blocks for many tumors. There is a renewed interest in therapeutically targeting glutamine metabolism due to the recent discovery of two novel glutaminase inhibitors. To quantify the dysregulation of the glutamate-glutamine equilibrium in breast cancer, metabolomics analysis of 270 clinical breast cancer samples and 97 normal breast samples was carried out using gas chromatography combined with time-of-flight mass spectrometry. Positive correlation between glutamate and glutamine in normal breast tissues switched to negative correlation between glutamate and glutamine in breast cancer tissues. Compared with the ratio of glutamate to glutamine in normal tissues, we found 56% of the ER1 tumor tissues and 88% of the ER2 tumor tissues glutamate-enriched. The glutamateto-glutamine ratio (GGR) significantly correlated with ER status (p 5 8.0E-09) and with tumor grade (p 5 3.3E-05). Higher levels of GGR were associated with prolonged overall survival in univariate analysis (HR 5 0.77, p 5 0.027) and in multivariate analysis (HR 5 0.73, p 5 0.038). GGR levels were reflected in an unsupervised clustering of metabolomics profiles. In a supervised analysis of metabolomics data and of genome-wide expression data, replacement of GGR by metabolite surrogate markers was feasible, while replacement of GGR by RNA markers had a limited accuracy. Functional analysis of the gene expression data showed negative correlation between glutamate enrichment and activation of peroxisome proliferator-activated receptor (PPAR) pathway. Our findings may have important implications for patient stratification related to utilization of glutaminase inhibitors.Altered energy metabolism is acknowledged as one of the emerging hallmarks of cancer. 1 Recently, there is a renewed interest in the metabolic transformation occurring in cancer cells and in targeting metabolic enzymes for cancer therapy. 2-4 However, the discovery of dysregulated metabolism in tumors dates back to the work of Otto Warburg in the 1920s. Warburg observed that cancer cells exhibit elevated glycolysis and that much of the generated pyruvate is fermented to lactate rather than feed to the TCA cycle and used for oxidative phosphorylation. 5,6 This phenomenon occurs under anaerobic and even under aerobic conditions ("aerobic glycolysis").A second change that occurs in central metabolism of many cancer cells is the alteration of glutamine metabolism. 7-9 Glutamine can be used to replenish the TCA cycle instead of glucose and feed in carbon and nitrogen for synthesis of nucleotides, several amino acids and glutathione. Further, glutamine can be used for energy production and in transformed cultured cells glutamine driven oxidative phosphorylation was recently shown to be the main source of

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Section: Discussionmentioning

confidence: 99%

Glutamate enrichment as new diagnostic opportunity in breast cancer

Budczies

Pfitzner

Győrffy

et al. 2014

Intl Journal of Cancer

110

View full text Add to dashboard Cite

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“…Markers of insulin resistance such as insulin level, insulin receptors, receptor for advanced glycation end products (RAGE), as well as acetylcholine and glutamate were also assessed using specific diagnostic kits (RayBiotech, USA), (cusabio biotech, USA), (Sigma-Aldrich Company, USA), and (Abnova, United Kingdom) [ 22 , 23 ].…”

Section: Methodsmentioning

confidence: 99%

Involvement of insulin resistance in D-galactose-induced age-related dementia in rats: Protective role of metformin and saxagliptin

Kenawy¹,

Hegazy²,

Hassan³

et al. 2017

PLoS ONE

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Age-related dementia is one of the most devastating disorders affecting the elderly. Recently, emerging data suggest that impaired insulin signaling is the major contributor in the development of Alzheimer’s dementia (AD), which is the most common type of senile dementia. In the present study, we investigated the potential therapeutic effects of metformin (Met) and saxagliptin (Saxa), as insulin sensitizing agents, in a rat model of brain aging and AD using D-galactose (D-gal, 150 mg/kg/day, s.c. for 90 successive days). Six groups of adult male Wistar rats were used: normal, D-gal, Met (500 mg/kg/day, p.o), and Saxa (1 mg/kg/day, p.o) control groups, as well as D-gal/Met and D-gal/Sax treated groups. Impaired learning and memory function was observed in rats treated with D-gal using Morris water maze test. Biochemical and histopathological findings also revealed some characteristic changes of AD in the brain that include the increased content of acetylcholine, glutamate, and phosphorelated tau, as well as deposition of amyloid plaques and neurofibrillary tangles. Induction of insulin resistance in experimentally aged rats was evidenced by increased blood glycated hemoglobin, brain contents of insulin and receptors for advanced glycated end-products, as well as decreased brain insulin receptor level. Elevation of oxidative stress markers and TNF-α brain content was also demonstrated. Met and Saxa, with a preference to Met, restored the normal memory and learning functions in rats, improved D-gal-induced state of insulin resistance, oxidative stress and inflammation, and ameliorated the AD biochemical and histopathological alterations in brain tissues. Our findings suggest that D-gal model of aging results in a diminishing of learning and memory function by producing a state of impaired insulin signaling that causes a cascade of deleterious events like oxidative stress, inflammation, and tau hyper-phosphorylation. Reversing of these harmful effects by the use of insulin-sensitizing drugs like Met and Saxa suggests their involvement in alleviation insulin resistance as the underlying pathology of AD and hence their potential use as anti-dementia drugs.

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“…The glutamate dehydrogenase activity was assayed by measuring NADH oxidation at 340 nm in 50 mmol of triethanolamine buffer containing 50 mmol of ammonium sulfate, 200 nmol of NADH, and 10 mmol of 2-oxoglutarate. Reactions were measured every 6 min (20). Protein concentrations were determined by the Bradford assay (4).…”

Section: Methodsmentioning

confidence: 99%

A New TetR Family Transcriptional Regulator Required for Morphogenesis inStreptomyces coelicolor

Hillerich

Westpheling

2008

J Bacteriol

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Both morphogenesis and antibiotic production in the streptomycetes are initiated in response to starvation, and these events are coupled. We previously described a transposon-generated mutant in Streptomyces coelicolor, SE293, that resulted in a bld strain that overproduced the antibiotic actinorhodin. The SCO1135 open reading frame identified by the insertion encodes a member of the TetR family of transcriptional regulators. Here we show that a constructed deletion of the SCO1135 open reading frame resulted in the same morphological and antibiotic production phenotype as the insertion mutant. The constructed deletion also resulted in constitutive expression of SCO1135 transcript, as well as that of the gene cluster immediately adjacent to it, SCO1134-1132, which encodes a putative molybdopterin binding complex. A His 6 -tagged version of the SCO1135 protein product was shown to bind the intergenic region between SCO1135 and SCO1134, which contains the apparent transcription start sites for each gene mapped by primer extension analysis. Increased expression of the SCO1134-1132 transcript in the SCO1135 deletion mutant also resulted in increased expression of xanthine dehydrogenase activity, confirming the predictions about these open reading framed based on protein similarity. We have designated the SCO1134-1142 gene cluster xdhABC and the regulator encoded by SCO1135 xdhR. We speculate that the inappropriate expression of xanthine dehydrogenase affects purine salvaging pathways at the onset of development, creating artificially high concentrations of both GTP and ppGpp and perturbing the pathways these molecules participate in for the initiation of morphogenesis and antibiotic production.

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A glutamate dehydrogenase-based method for the assay ofl-glutamic acid: Formation of pyridine nucleotide fluorescent derivatives

Cited by 20 publications

References 13 publications

Glutamate enrichment as new diagnostic opportunity in breast cancer

Glutamate enrichment as new diagnostic opportunity in breast cancer

Involvement of insulin resistance in D-galactose-induced age-related dementia in rats: Protective role of metformin and saxagliptin

A New TetR Family Transcriptional Regulator Required for Morphogenesis inStreptomyces coelicolor

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