Regulation of glutamate synthase from Bacillussubtilis by Glutamine

The Bacillus subtilis gltAB operon, encoding glutamate synthase, requires a specific positive regulator, GltC, for its expression and is repressed by the global regulatory protein TnrA. The factor that controls TnrA activity, a complex of glutamine synthetase and a feedback inhibitor, such as glutamine, is known, but the signal for modulation of GltC activity has remained elusive. GltC-dependent gltAB expression was drastically reduced when cells were grown in media containing arginine or ornithine or proline, all of which are inducers and substrates of the Roc catabolic pathway. Analysis of gltAB expression in mutants with various defects in the Roc pathway indicated that rocG-encoded glutamate dehydrogenase was required for such repression, suggesting that the substrates or products of this enzyme are the real effectors of GltC. Given that RocG is an enzyme of glutamate catabolism, the main regulatory role of GltC may be prevention of a futile cycle of glutamate synthesis and degradation in the presence of arginine-related amino acids or proline. In addition, high activity of glutamate dehydrogenase was incompatible with activity of TnrA.

Section: Resultsmentioning

confidence: 99%

Modulation of Activity of Bacillus subtilis Regulatory Proteins GltC and TnrA by Glutamate Dehydrogenase

Belitsky

Sonenshein

2004

“…A longer time was required to filter cells grown with glutamate rather than NH4 plus glutamate; however, essentially the same amino acid profile is observed in samples that remain on the filter for 60 s. After 20 min the solution and filter were placed in a Waring blender for 1 min. The suspension was clarified by centrifugation (8,000 x g) at 4°C, perchlorate was precipitated by the addition of 8 ml of 0.2 M K2CO3 to give a final pH of 6.5 to 7.0, KCl03 was removed by centrifugation, and the supernatant was assayed for amino acids and NH4 (5,12). Purification of GS.…”

Section: Methodsmentioning

confidence: 99%

“…Recently, we reported that the activity of GOGAT in B. subtilis is inversely proportional to the internal concentration ofglutamine rather than of glutamate (5). In fact, the intracellular glutamine concentration is significantly greater in cells grown with glutamate than in cells grown with NH' or NHW plus glutamate.…”

mentioning

confidence: 91%

“…In several gram-negative bacteria (1,17,18) and Bacillus species (17,18,20), GOGAT activity decreases in cells grown with glutamate as a nitrogen source and increases as a result of the addition of NW to the growth medium. Al- though GOGAT appears to be regulated in a negative manner by glutamate, the intracellular glutamate concentration remains essentially unchanged in strains cultured in different growth conditions (5,17).…”

mentioning

confidence: 99%

See 1 more Smart Citation

Effect of glutamine on enzymes of nitrogen metabolism in Bacillus subtilis

Deshpande¹,

Katze²,

Kane³

1981

An earlier study of the regulation of glutamate synthase (GOGAT) in Bacillus subtilis (Deshpande et al., Bichem. Biophys. Res. Commun. 95:55--60, 1980) revealed an inverse relationship between the specific activity of this essential ammonia-assimilatory enzyme and the intracellular pool of glutamine: GOGAT activity decreased when the internal glutamine concentration reached or exceeded 2.5 mM. This finding prompted the present investigation of the intracellular events linking glutamine formation to the regulation of GOGAT. A growing culture of B. subtilis was shifted from glutamate plus NH+4 medium (high GOGAT activity) to glutamate medium (low GOGAT activity). At various times after the shift, the intracellular concentrations of aspartate, glutamate, glutamine, alanine, and NH+4 and the activities of GOGAT and glutamine synthetase (GS) were measured. After 30 min, the only significant pool level change was an eightfold increase in glutamine, which paralleled a 2- to 3-fold increase in GS activity. Approximately 15 min after the glutamine pool reached its peak, GOGAT activity began to decrease and eventually declined 2.5-fold. In contrast, when B. subtilis was shifted from glutamate medium to glutamate plus NH+4 medium, there was a 1- to 2-h lag before the glutamine pool and GS activity approached a steady state. As a result, GOGAT activity was low until the concentration of glutamine dropped below 2.5 mM. We propose that glutamine is an important regulatory element in the control of GOGAT activity and that one form of GOGAT regulation involves enzyme inactivation. In addition, these results indicate that glutamine is neither a corepressor nor a feedback inhibitor of GS.

“…When extracts of strains SU1 and SU4 were mixed, 70% of the GOGAT activity of the parental strains was observed so that if GOGAT of P. aeruginosa resembles that of other organisms in comprising two nonidentical subunits (18,29), it seems likely that the mutations affect a different GOGAT subunit in each strain. The lesion in strain US1 on August 6, 2020 by guest http://jb.asm.org/ Downloaded from has manifold effects, and preliminary experiments indicate that intracellular concentrations of glutamine in this mutant are much higher during growth on all media than in the parent strain; there is some evidence that glutamine represses GOGAT synthesis in B. subtilis (8) and that this may account for the lack of GO-GAT activity in strain US1. An investigation of the levels of enzymes and intermediary metabolites in partial revertants of strain US1 is currently being directed towards a more detailed characterization of this strain.…”

Section: Isolation Of Mutants Gogat-negative Mutantsmentioning

confidence: 97%

Growth of Pseudomonas aeruginosa mutants lacking glutamate synthase activity

Brown¹,

Tata²

1981

Mutant strains SU1, SU4, and US1 lacking glutamate synthase (GOGAT) activity were isolated from strains of P. aeruginosa for which histidine is a growth rate-limiting source of nitrogen. Strains SUl and SU4 were unable to grow when a low concentration of ammonia and a variety of compounds, including histidine, were supplied as sole sources of nitrogen. A revertant of strain SU1, strain 39, produced no GOGAT but high levels of nicotinamide adenine dinucleotidedependent glutamate dehydrogenase and had restored ability to grow on a limited number of nitrogen sources. Strain US1 grew at the same rate in histidine medium as did its parent; it was derepressed for glutamine synthase synthesis, and histidase was less sensitive to repression by ammonia than in the parent strain. We conclude that GOGAT is not essential for growth on histidine but high levels of glutamine synthase are required and high levels of nicotinamide adenine dinucleotide-dependent glutamate dehydrogenase can sustain growth at low concentrations of ammonia in the absence of GOGAT.