TheN‐acetylglutamate synthase/N‐acetylglutamate kinase metabolon ofSaccharomyces cerevisiaeallows co‐ordinated feedback regulation of the first two steps in arginine biosynthesis

Pauwels, Kris; Abadjieva, Agnes; Hilven, Pierre; Stankiewicz, Anna; Crabeel, Marjolaine

doi:10.1046/j.1432-1033.2003.03477.x

Cited by 43 publications

(42 citation statements)

References 45 publications

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“…71,2007 SURPRISING ARGININE BIOSYNTHESIS 41 In the yeast Saccharomyces cerevisiae, both NAGS and NAGK are inhibited by arginine, but NAGS must be associated with NAGK to remain both functional and sensitive to arginine. Deletion of the NAGS gene decreases the sensitivity of the kinase, while making the kinase insensitive renders NAGS insensitive as well (1,56). The situation appears to be similar in Neurospora crassa (83).…”

Section: Feedback Inhibition Of Ornithine Synthesismentioning

confidence: 85%

Surprising Arginine Biosynthesis: a Reappraisal of the Enzymology and Evolution of the Pathway in Microorganisms

Labedan

Glansdorff

2007

Microbiol Mol Biol Rev

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SUMMARY Major aspects of the pathway of de novo arginine biosynthesis via acetylated intermediates in microorganisms must be revised in light of recent enzymatic and genomic investigations. The enzyme N-acetylglutamate synthase (NAGS), which used to be considered responsible for the first committed step of the pathway, is present in a limited number of bacterial phyla only and is absent from Archaea. In many Bacteria, shorter proteins related to the Gcn5-related N-acetyltransferase family appear to acetylate l-glutamate; some are clearly similar to the C-terminal, acetyl-coenzyme A (CoA) binding domain of classical NAGS, while others are more distantly related. Short NAGSs can be single gene products, as in Mycobacterium spp. and Thermus spp., or fused to the enzyme catalyzing the last step of the pathway (argininosuccinase), as in members of the Alteromonas-Vibrio group. How these proteins bind glutamate remains to be determined. In some Bacteria, a bifunctional ornithine acetyltransferase (i.e., using both acetylornithine and acetyl-CoA as donors of the acetyl group) accounts for glutamate acetylation. In many Archaea, the enzyme responsible for glutamate acetylation remains elusive, but possible connections with a novel lysine biosynthetic pathway arose recently from genomic investigations. In some Proteobacteria (notably Xanthomonadaceae) and Bacteroidetes, the carbamoylation step of the pathway appears to involve N-acetylornithine or N-succinylornithine rather than ornithine. The product N-acetylcitrulline is deacetylated by an enzyme that is also involved in the provision of ornithine from acetylornithine; this is an important metabolic function, as ornithine itself can become essential as a source of other metabolites. This review insists on the biochemical and evolutionary implications of these findings.

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Section: Feedback Inhibition Of Ornithine Synthesismentioning

confidence: 85%

Surprising Arginine Biosynthesis: a Reappraisal of the Enzymology and Evolution of the Pathway in Microorganisms

Labedan

Glansdorff

2007

Microbiol Mol Biol Rev

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“…ARG5,-6 encode the enzymes that catalyze the second and third steps and are translated into a pre-protein that is imported into mitochondria, where it is cleaved, resulting in separate proteins, i.e., N-acetylglutamate kinase (Arg6) and N-acetylglutamyl-phosphate reductase (Arg5) (Boonchird et al 1991). The first two enzymes in the pathway, N-acetylglutamate synthase (Arg2) and N-acetylglutamate kinase (Arg6), bind each other, forming a complex that is necessary for their stability and for feedback inhibition by arginine (Abadjieva et al 2000(Abadjieva et al , 2001Pauwels et al 2003). The ornithine synthesized in mitochondria is transported to the cytoplasm via the mitochondrial carrier protein Ort1 (Table 5), and the remaining steps are carried out in the cytoplasm.…”

Section: Pathway Genesmentioning

confidence: 99%

Regulation of Amino Acid, Nucleotide, and Phosphate Metabolism in Saccharomyces cerevisiae

2012

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Ever since the beginning of biochemical analysis, yeast has been a pioneering model for studying the regulation of eukaryotic metabolism. During the last three decades, the combination of powerful yeast genetics and genome-wide approaches has led to a more integrated view of metabolic regulation. Multiple layers of regulation, from suprapathway control to individual gene responses, have been discovered. Constitutive and dedicated systems that are critical in sensing of the intra- and extracellular environment have been identified, and there is a growing awareness of their involvement in the highly regulated intracellular compartmentalization of proteins and metabolites. This review focuses on recent developments in the field of amino acid, nucleotide, and phosphate metabolism and provides illustrative examples of how yeast cells combine a variety of mechanisms to achieve coordinated regulation of multiple metabolic pathways. Importantly, common schemes have emerged, which reveal mechanisms conserved among various pathways, such as those involved in metabolite sensing and transcriptional regulation by noncoding RNAs or by metabolic intermediates. Thanks to the remarkable sophistication offered by the yeast experimental system, a picture of the intimate connections between the metabolomic and the transcriptome is becoming clear.

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“…The importance of the controlling function of the arginine-inhibitable form is highlighted by the observation that in photosynthetic organisms, NAGK is a key target of the nitrogen signaling protein P II : when nitrogen is abundant, P II binds to NAGK, relieving arginine inhibition and allowing fast arginine synthesis (4,13,16,23). Further controlling functions for arginine-sensitive NAGKs have been unveiled in Saccharomyces cerevisiae, where NAGK is involved in gene regulation (12) and in the formation of a complex with NAGS having the characteristics of a true metabolon, in which the regulatory properties of NAGS and NAGK are influenced mutually (1,17,18).In an effort to ascertain the bases for the arginine sensitivity of NAGKs, we determined the crystal structures of four NAGKs, with one of them, that from E. coli (EcNAGK) (9, 19), being arginine insensitive, and the other three, from P. aeruginosa (PaNAGK), Thermotoga maritima (TmNAGK) (20), and Synechococcus elongatus (bound to P II ) (16), being inhibited by arginine. These studies have revealed that EcNAGK is homodimeric and that arginine-sensitive NAGKs are hexameric and composed of three homodimers, with each one of them resembling EcNAGK (20).…”

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confidence: 99%

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Basis of Arginine Sensitivity of Microbial N -Acetyl- l -Glutamate Kinases: Mutagenesis and Protein Engineering Study with the Pseudomonas aeruginosa and Escherichia coli Enzymes

Fernández-Murga

Rubio

2008

J Bacteriol

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N-Acetylglutamate kinase (NAGK) catalyzes the second step of arginine biosynthesis. In Pseudomonas aeruginosa, but not in Escherichia coli, this step is rate limiting and feedback and sigmoidally inhibited by arginine. Crystal structures revealed that arginine-insensitive E. coli NAGK (EcNAGK) is homodimeric, whereas arginine-inhibitable NAGKs, including P. aeruginosa NAGK (PaNAGK), are hexamers in which an extra N-terminal kinked helix (N-helix) interlinks three dimers. By introducing single amino acid replacements in PaNAGK, we prove the functionality of the structurally identified arginine site, as arginine site mutations selectively decreased the apparent affinity for arginine. N-helix mutations affecting R24 and E17 increased and decreased, respectively, the apparent affinity of PaNAGK for arginine, as predicted from enzyme structures that revealed the respective formation by these residues of bonds favoring inaccessible and accessible arginine site conformations. N-helix N-terminal deletions spanning >16 residues dissociated PaNAGK to active dimers, those of <20 residues decreased the apparent affinity for arginine, and complete N-helix deletion (26 residues) abolished arginine inhibition. Upon attachment of the PaNAGK N-terminal extension to the EcNAGK N terminus, EcNAGK remained dimeric and arginine insensitive. We concluded that the N-helix and its C-terminal portion after the kink are essential but not sufficient for hexamer formation and arginine inhibition, respectively; that the N-helix modulates NAGK affinity for arginine and mediates signal transmission between arginine sites, thus establishing sigmoidal arginine inhibition kinetics; that the mobile ␣H-␤16 loop of the arginine site is the modulatory signal receiver; and that the hexameric architecture is not essential for arginine inhibition but is functionally essential for physiologically relevant arginine control of NAGK.Microorganisms and plants make arginine from glutamate in eight steps (6), among which the first five (glutamate3N-acetylglutamate3N-acetylglutamyl-␥-phosphate3N-acetylglutamate-␥-semialdehyde3N-acetylornithine3ornithine) involve N-acetylated intermediates and generate the arginine precursor ornithine (5,21,22). The absence of steps 2 to 5 in animals (2, 14) renders the enzymes catalyzing these steps potential candidates for developing antibacterials and biocides. In some microorganisms, such as Escherichia coli, the arginine biosynthetic route is linear because ornithine is produced by the hydrolysis of N-acetylornithine (5). In such cases, the first enzyme of the route, N-acetylglutamate synthase (NAGS), catalyzes the controlling step and is feedback inhibited by the final product, arginine. Many more organisms, including yeasts, algae, plants, and many bacteria, such as Pseudomonas aeruginosa, have a more evolved, cyclic variant of this route in which the acetyl group is recycled by its transfer from acetylornithine to glutamate (5,6,22). In this case, NAGS plays a purely anaplerotic role, and the main controlling step that ...

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TheN‐acetylglutamate synthase/N‐acetylglutamate kinase metabolon ofSaccharomyces cerevisiaeallows co‐ordinated feedback regulation of the first two steps in arginine biosynthesis

Cited by 43 publications

References 45 publications

Surprising Arginine Biosynthesis: a Reappraisal of the Enzymology and Evolution of the Pathway in Microorganisms

Surprising Arginine Biosynthesis: a Reappraisal of the Enzymology and Evolution of the Pathway in Microorganisms

Regulation of Amino Acid, Nucleotide, and Phosphate Metabolism in Saccharomyces cerevisiae

Basis of Arginine Sensitivity of Microbial N -Acetyl- l -Glutamate Kinases: Mutagenesis and Protein Engineering Study with the Pseudomonas aeruginosa and Escherichia coli Enzymes

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