Regulation of glutamine synthetase activity in the unicellular cyanobacterium Synechocystis sp. strain PCC 6803 by the nitrogen source: effect of ammonium

Abstract

Glutamine synthetase activity from Synechocystis sp. strain PCC 6803 is regulated as a function of the nitrogen source available in the medium. Addition of 0.25 mM NH4Cl to nitrate-grown cells promotes a clear short-term inactivation of glutamine synthetase, whose enzyme activity decreases to 5 to 10% of the initial value in 25 min. The intracellular levels of glutamine, determined under various conditions, taken together with the results obtained with azaserine (an inhibitor of transamidases), rule out the possibility that glutamine per se is responsible for glutamine synthetase inactivation. Nitrogen starvation attenuates the ammonium-mediated glutamine synthetase inactivation, indicating that glutamine synthetase regulation is modulated through the internal balance between carbon-nitrogen compounds and carbon compounds. The parallelism observed between the glutamine synthetase activity and the internal concentration of a-ketoglutarate suggests that this metabolite could play a role as a positive effector of glutamine synthetase activity in Synechocystis sp. Despite the similarities of this physiological system to that described for enterobacteria, the lack of in vivo 32p labeling of glutamine synthetase during the inactivation process excludes the existence of an adenylylation-deadenylylation system in this cyanobacterium.Glutamine synthetase (GS; EC 6.3.1.2) plays a central role in nitrogen metabolism by prokaryotes. This central role requires that both the synthesis and activity of this enzyme be strictly regulated in response to the available nitrogen source. In enterobacteria this regulation takes place at two different levels, first by modulating enzyme activity by an adenylylation-deadenylylation system (35) and second by regulating GS synthesis through transcription from different promoters (18). The adenylylation of GS makes it more susceptible to a cumulative feedback inhibition by metabolites derived from ammonium assimilation, such as amino acids or nucleotides (35). In contrast, most of the grampositive bacteria lack this system, and the regulation of the enzyme appears to involve a single feedback mechanism, in which GS is strongly inhibited by glutamine (5,14).In cyanobacteria, ammonium assimilation takes place mainly by the sequential action of GS and glutamate synthase (GOGAT) (GS-GOGAT pathway) (21). The regulation of GS synthesis in Anabaena sp. strain PCC 7120 has been well characterized; several promoters have been found for the structural GS gene, ginA (37). Selective transcription from these promoters determines that in ammonium-grown cells the level of GS protein is about 50% lower than that observed under dinitrogen-fixing conditions (37). Recently, we have reported a repression of GS synthesis by ammonium in the cyanobacteria Synechocystis sp. and Calothrix sp. (22).The existence of an adenylylation-deadenylylation system, as observed in the majority of gram-negative bacteria, has been previously discounted for the genus Anabaena (8), and it has been proposed that regulation of GS act...

Cell surface acid-base properties of the cyanobacterium Synechococcus : Influences of nitrogen source, growth phase and N:P ratios

“…As a result, the numerical data from the heat-killed titrations has the sole purpose of testing whether microbial biomass undergoes changes in acid-base behavior as the result of different growth conditions (which a key objective of this study). (Mérida et al, 1991). This implies that the reduced ligand concentrations inferred for the live nitrate-reducing cultures are a byproduct of cell wall compositional changes associated with growth by that metabolism.…”

Purification and Characterization of A New Type of Glutamine Synthetase from Cyanobacteria

“…GS transferase activity was determined either in vitro or in situ by the formation of y-glutamylhydroxamate, as previously described [26]. In nondenaturating PAGE, GS transferase activity was determined as follows.…”

Decoupling of ammonium regulation and ntcA transcription in the diazotrophic marine cyanobacterium Trichodesmium sp. IMS101

“…In the presence of high concentrations of ammonium, cyanobacteria are neither capable of assimilating other N sources, nor can they engage in N 2 fixation and heterocyst development . Type I glutamine synthetase activities are repressed by ammonium additions (Merida et al, 1991). Ammonium reversibly inhibits nitrate assimilation (Ohmori et al, 1977;Flores et al, 1980).…”