Ammonium Assimilation by Candida albicans and Other Yeasts: Evidence for Activity of Glutamate Synthase

Holmes, Ann R.; Collings, A.; Farnden, Kevin J. F.; Shepherd, Maxwell G.

doi:10.1099/00221287-135-6-1423

Cited by 29 publications

(18 citation statements)

References 20 publications

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“…This novel report of an allosteric 2-oxoglutarate interaction with NADP-GDH appears to be a regulatory feature common at least to the four aspergilli tested. Among the fungal enzymes studied so far, only yeast NADP-GDHs are reported to exhibit sigmoidal 2-oxoglutarate saturation (Holmes et al, 1989;Perysinakis et al, 1994;DeLuna et al, 2001). The two S. cerevisiae isozymes (Gdh1p and Gdh3p) are implicated in fine-tuning yeast glutamate synthesis and carbon metabolism during the diauxic shift.…”

Section: Discussionmentioning

confidence: 99%

“…So far, only yeast NADP-GDHs are known to show a co-operative response towards 2-oxoglutarate (Holmes et al, 1989;Perysinakis et al, 1994;DeLuna et al, 2001). As a step towards understanding the role of NADP-GDH in metabolic regulation at the carbon-nitrogen interface we have (i) devised an effective method to purify Abbreviations: GDH, glutamate dehydrogenase; TCA, tricarboxylic acid.…”

Section: Introductionmentioning

confidence: 99%

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Allosteric NADP-glutamate dehydrogenase from aspergilli: purification, characterization and implications for metabolic regulation at the carbon–nitrogen interface

Noor

Punekar

2005

Microbiology

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NADP-dependent glutamate dehydrogenase (NADP-GDH) mediates fungal ammonium assimilation through reductive synthesis of glutamate from 2-oxoglutarate. By virtue of its position at the interface of carbon and nitrogen metabolism, biosynthetic NADP-GDH is a potential candidate for metabolic control. In order to facilitate characterization, a new and effective dye-affinity method was devised to purify NADP-GDH from two aspergilli, Aspergillus niger and Aspergillus nidulans. The A. niger NADP-GDH was characterized at length and its kinetic interaction constants with glutamate (K m 34?7 mM) and ammonium (K m 1?05 mM; K i 0?4 mM) were consistent with an anabolic role. Isophthalate, 2-methyleneglutarate and 2,4-pyridinedicarboxylate were significant inhibitors, with respective K i values of 6?9, 9?2 and 202?0 mM. The A. niger enzyme showed allosteric properties and a sigmoid response (n H =2?5) towards 2-oxoglutarate saturation. The co-operative behaviour was a feature common to NADP-GDH from Aspergillus awamori, A. nidulans and Aspergillus oryzae. NADP-GDH may therefore be a crucial determinant in adjusting 2-oxoglutarate flux between the tricarboxylic acid cycle and glutamate biosynthesis in aspergilli.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Allosteric NADP-glutamate dehydrogenase from aspergilli: purification, characterization and implications for metabolic regulation at the carbon–nitrogen interface

Noor

Punekar

2005

Microbiology

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“…In studies on the primary nitrogen metabolism of fungi, yeasts and ascomycetes have been investigated preferentially (Brown et al, 1974;Casper et al, 1985;Dunn-Coleman et al, 198 1 ;Holmes et al, 1989;Kusnan et al, 1987Kusnan et al, ,1989Limon-Lason et al, 1977). In this paper the primary nitrogen metabolism of the basidomycetic fungus Stropharia semiglobata was investigated.…”

Section: Introductionmentioning

confidence: 99%

The involvement of glutamate dehydrogenase and glutamine synthetase/glutamate synthase in ammonia assimilation by the basidiomycete fungus Stropharia semiglobata

Schwartz

Misri

Fock

1991

Journal of General Microbiology

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~When grown on ammonia as sole nitrogen source, the basidiomycete fungus Stropharia semigfobatu assimilated nitrogen via glutamate dehydrogenase (GDH). The two isoenzymes of GDH were both repressed by increasing concentrations of ammonia, but NADP-GDH (EC 1.4.1.4) was far more active than NAD-GDH (EC 1.4.1.2). Glutamine synthetase (GS; EC 6.3.1.2) and glutamate synthase (GOGAT; EC 1.4.7.1) were also active in S. semigfobatu and these enzyme activities were independent of the ammonia concentration in the suspension. From enzyme activity and lSN-labelling studies of amino acids, it is concluded that both GDH and GS/GOGAT contribute to ammonia assimilation in this fungus.

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“…Y-to-M transition studies were also carried out with inhibitors such as cycloheximide (a protein synthesis inhibitor), isophthalic acid (a strong competitive NAD-GDH inhibitor) (14), methionine sulfoximine (an irreversible GS inhibitor) (11), and azaserine, a glutamine analog (a GOGAT inhibitor) (4).…”

Section: Resultsmentioning

confidence: 99%

Significance of NADP/NAD glutamate dehydrogenase ratio in the dimorphic behavior of Benjaminiella poitrasii and its morphological mutants

1992

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Studies on the levels of glutamate dehydrogenase (GDH), glutamine synthetase, and glutamate synthase were carried out as a function of temperature, nutritional conditions, and the morphological (yeast or mycelium) form of Benjaminiella poitrasii. Since both NAD-and NADP-dependent GDH activities were found in B. poitrasii, the quantitative relation between these two enzymes expressed as the NADP-GDH/NAD-GDH activity ratio (GDH ratio) was studied to evaluate its possible role in the morphogenesis. In the yeast-to-mycelium transition, a decrease in the GDH ratio occurred (between 1 and 2 h) and germ tube formation could be observed only at 3 h. Under similar sets of experimental conditions, exogenous addition 1.0 mM of a-ketoglutarate delayed germ tube emergence (4 h) compared with the control. On the other hand, in the presence of 1.0 mM glutamate an earlier onset of the germ tube formation was noted. The morphological (monomorphic) mutants, Y-2 and Y-5, showed a high GDH ratio and maintained the yeast morphology.The dimorphic zygomycete Benjaminiella poitrasii has the potential to differentiate into alternative morphologies depending on growth temperature and/or nutritional conditions. The organism grows in mycelial form (M form) at 28°C whereas development of the yeast form (Y form) occurs at 37°C. In addition, optimal yeast growth requires the presence of a hexose as a carbon source as well as an organic amino compound as a nitrogen source. In contrast, the mycelial growth occurs on a variety of carbon sources and with ammonium salts as the sole nitrogen source (5). These observations suggest differential expression of some key enzymes of carbon and nitrogen metabolism in the two morphological forms. Generally, in fungi, the carbon and nitrogen metabolisms are connected via ammonia assimilation and the participating enzymes are NAD-dependent (EC 1.4.1.2) and NADP-dependent (EC 1.4.1.4) glutamate dehydrogenase (GDH), glutamate synthase (GOGAT; EC 1.4.1.13), and glutamine synthetase (GS; EC 6.3.1.2) (8).Since the dimorphic (Y-to-M) transitions in B. poitrasii can be regulated by the carbon-nitrogen balance, the changes in the levels of the aforementioned enzymes were studied to evaluate their possible role. Similar studies were also carried out with monomorphic Y form mutants (Y-2 and Y-5) isolated after N-methyl-N'-nitro-N-nitrosoguanidine treatment of parent strain spores (5). (pH 7.2) was prepared. The pH was adjusted to 7.2 with 2 M NaOH or 1.0 M HCI. The reaction mixture (1 ml) containing 0.4 ml of the above assay mixture, 0.2 ml of crude extract, and 0.35 ml of glass-distilled water was preincubated for 5 min at 37°C. Then the reaction was initiated by the addition MATERUILS AND METHODS

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Ammonium Assimilation by Candida albicans and Other Yeasts: Evidence for Activity of Glutamate Synthase

Cited by 29 publications

References 20 publications

Allosteric NADP-glutamate dehydrogenase from aspergilli: purification, characterization and implications for metabolic regulation at the carbon–nitrogen interface

Allosteric NADP-glutamate dehydrogenase from aspergilli: purification, characterization and implications for metabolic regulation at the carbon–nitrogen interface

The involvement of glutamate dehydrogenase and glutamine synthetase/glutamate synthase in ammonia assimilation by the basidiomycete fungus Stropharia semiglobata

Significance of NADP/NAD glutamate dehydrogenase ratio in the dimorphic behavior of Benjaminiella poitrasii and its morphological mutants

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