NAD and NADP L-Glutamate Dehydrogenase Activity and Ammonium Regulation in Aspergillus nidulans

Kinghorn, James R.; Pateman, John

doi:10.1099/00221287-78-1-39

Cited by 98 publications

(61 citation statements)

References 16 publications

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“…The biosynthetic role of fungal NADP-GDH is well supported by genetic (Fincham, 1962;Kinghorn & Pateman, 1973;Fincham et al, 2000), physiological (Martin et al, 1988;Kusnan et al, 1989;Schwartz et al, 1991) and kinetic (Hudson & Daniel, 1993) data. The kinetic constants obtained for the various substrates of the A. niger NADP-GDH (Table 2) are consistent with its role in ammonium assimilation.…”

Section: Discussionmentioning

confidence: 83%

“…A convenient purification method, a prerequisite in elucidating such features, was therefore first established. The specific activity of NADP-GDH is often influenced by the nature of the growth medium (Kinghorn & Pateman, 1973;Cardoza et al, 1998;Pedersen et al, 1999). The A. niger NADP-GDH activity profile, when resolved on DEAESephacel matrix, was qualitatively identical irrespective of the nature of the nitrogen source used for cell growth (data not shown).…”

Section: Resultsmentioning

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: 83%

Section: Resultsmentioning

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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“…The phenotypes of mutants affected in ammonium assimilation indicate that ammonium alone is not the key effector for the modulation of AreA function. gdhA mutants are partially derepressed on ammonium, suggesting that ammonium must be metabolized, possibly to glutamine, to generate the signal(s) for full nitrogen metabolite repression (4,21,22). Previous studies of glnA mutants in A. nidulans and gln-1 mutants in Neurospora crassa have presented conflicting evidence about the role of glutamine and have even led to the suggestion that the glutamine synthetase protein itself may have a regulatory role (10,15,16,17,28,37,38).…”

mentioning

confidence: 99%

“…The major route of ammonium assimilation in Aspergillus nidulans is through the action of NADP-linked glutamate dehydrogenase (NADP-GDH), encoded by the gdhA gene, to yield glutamate (4,22,23). In addition, ammonium can be assimilated to glutamate by the combined action of glutamine synthetase (GS) and glutamate synthase (GOGAT).…”

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confidence: 99%

Role of Glutamine Synthetase in Nitrogen Metabolite Repression in Aspergillus nidulans

et al. 2001

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Glutamine synthetase (GS), EC 6.3.1.2, is a central enzyme in the assimilation of nitrogen and the biosynthesis of glutamine. We have isolated the Aspergillus nidulans glnA gene encoding GS and have shown that glnA encodes a highly expressed but not highly regulated mRNA. Inactivation of glnA results in an absolute glutamine requirement, indicating that GS is responsible for the synthesis of this essential amino acid. Even when supplemented with high levels of glutamine, strains lacking a functional glnA gene have an inhibited morphology, and a wide range of compounds have been shown to interfere with repair of the glutamine auxotrophy. Heterologous expression of the prokaryotic Anabaena glnA gene from the A. nidulans alcA promoter allowed full complementation of the A. nidulans glnA⌬ mutation. However, the A. nidulans fluG gene, which encodes a protein with similarity to prokaryotic GS, did not replace A. nidulans glnA function when similarly expressed. Our studies with the glnA⌬ mutant confirm that glutamine, and not GS, is the key effector of nitrogen metabolite repression. Additionally, ammonium and its immediate product glutamate may also act directly to signal nitrogen sufficiency.

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“…The properties of various areA mutants indicate that the transcriptional activation function of the AreA protein is modulated by changes in the nitrogen status of the cells perceived through interactions involving at least the C-terminal regions of the protein (4,30). A number of other genes have been identified as influencing, directly or indirectly, the nitrogen regulatory circuit, including tamA, meaA, meaB, gdhA, sarA, and sarB (7,8,22,23,27). Of these, the tamA gene has been the most controversial.…”

mentioning

confidence: 99%

The tamA gene of Aspergillus nidulans contains a putative zinc cluster motif which is not required for gene function

et al. 1996

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Expression of many nitrogen catabolic enzymes is controlled by nitrogen metabolite repression in Aspergillus nidulans. Although the phenotypes of tamA mutants have implicated this gene in nitrogen regulation, its function is unknown. We have cloned the tamA gene by complementation of a new tamA allele. The tamA sequence shares significant homology with the UGA35/DAL81/DURL gene of Saccharomyces cerevisiae. In vitro mutagenesis of sequences encoding a putative zinc cluster DNA binding domain indicated that this motif is not required for in vivo TamA function.

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NAD and NADP L-Glutamate Dehydrogenase Activity and Ammonium Regulation in Aspergillus nidulans

Cited by 98 publications

References 16 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

Role of Glutamine Synthetase in Nitrogen Metabolite Repression in Aspergillus nidulans

The tamA gene of Aspergillus nidulans contains a putative zinc cluster motif which is not required for gene function

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