Soluble glutamine synthetase activity (L-glutamate:ammonia ligase, ADP forming, EC 6.3.1.2) was purified to electrophoretic homogeneity from the filamentous non-N2-fixing cyanobacterium Phormidium laminosum (OH-1-p.C1l) by using conventional purification procedures in the absence of stabilizing ligands. The pure enzyme showed a specific activity of 152 pumol of y-glutamylhydroxamate formed * min-(transferase activity), which corresponded to 4.4 pumol of P1 released min-' (biosynthetic activity). (20), photosynthetic bacteria (3), and a wide range of cyanobacteria (see reference 11 for a recent review). However, in addition to the GS-glutamate synthase cycle, glutamate dehydrogenase (EC 1.4.1.4) (16) and alanine dehydrogenase (EC 1.4.1.1) (24) may also contribute to ammonium assimilation in cyanobacteria, especially under certain nutritional conditions of nitrogen availability. Recently, an alternative route to GSglutamate synthase for the primary assimilation of ammonium in Anabaena sp. strain 1F, involving the participation of GS in collaboration with carbamylphosphate synthetase (EC 2.7.2.9), has been proposed (5). Consequently, GS is subject to intensive regulatory control.Although the GS isolated from bacterial sources are similar in a number of features, such as molecular size (about 600 kilodaltons [kDa]), subunit structure (dodecamers composed of identical subunits of about 50 kDa), and the requirement of divalent cations for activity, their regulatory properties appear to be quite different. For example, the enzyme from Escherichia coli (28) and several other gramnegative bacteria (13) is regulated by covalent modification through adenylylation-deadenylylation. In contrast, the GS from gram-positive bacteria is susceptible to feedback regu-* Corresponding author.lation by intracellular concentrations of adenine nucleotides, amino acids, and divalent cations rather than by covalent modification (17). However, in blue-green algae (cyanobacteria), the enzyme can be activated by dithiol compounds in vitro or by reduced thioredoxin (27). The cyanobacterial enzyme is not susceptible to regulation by covalent modification, and the availability of divalent cations and presence of feedback inhibitors (mainly Gly, L-Ala, and L-Ser) may play the dominant role in regulating GS in vivo (22,30). Moreover, the cyanobacterial enzyme shows cumulative inhibition with mixtures of inhibitors (i.e., AMP, L-Ser, and L-Asp), which indicates that there are independent binding sites on the enzyme for the inhibitors (30). The Anabaena flos-aquae GS was also found to be regulated by the energy charge of the cell, and cooperative inhibition was caused by CTP and any single nucleotide (18).GS from several species of the filamentous N2-fixing cyanobacterial genera Anabaena (18,23,25,29) and Nostoc (25) has been purified to electrophoretical homogeneity and characterized. Despite abundant information on the enzyme from N2-fixing cyanobacteria, only data regarding the GS from the unicellular non-N2-fixing cyanobacterium Anacystis nidu...