Cyanobacteria assimilate carbon dioxide through the Calvin cycle and therefore must regulate the activity of ribulose 1,5-bisophosphate carboxylase. Using an in situ assay, as well as measuring the activity in crude, partially purified, and homogeneous preparations, we can show that a number of phosphorylated intermediates exert a regulatory role. Three diverse organisms, Agmenellum quadruplicatum, Aphanocapsa 6714, and Anabaena sp. CA, were studied, and it was found that the in situ and cell-free carboxylase activities were particularly affected by low levels ofphosphogluconate and reduced nicotinamide adenine dinucleotide phosphate. There was a marked activation by these ligands when the inactive enzyme was assayed in the presence of low levels of bicarbonate, a result significantly different from a previous report. Moreover, the fully activated enzyme was inhibited by phosphogluconate. In situ Anabaena CA carboxylase activity exhibited a particular capacity for activation by phosphogluconate and reduced nicotinamide adenine dinucleotide phosphate. However, activation of the crude, partially purified, or homogeneous Anabaena CA carboxylase by phosphogluconate and reduced nicotinamide adenine dinucleotide phosphate was significantly decreased when compared with enzyme activity in permeabilized cells. It appears that the microenvironment or the conformation of the enzyme within the cell may be significantly different from that of the isolated enzyme.Cyanobacteria (blue-green algae) form organic matter by assimilating carbon dioxide. The enzyme which catalyzes this primary CO2 assimilation is ribulose 1,5-bisphosphate (Rbu-P2) carboxylase (3-phospho-D-glycerate carboxylase [dimerizing]; EC 4.1.1.39). Various phosphorylated intermediates, particularly those characteristic of photosynthetic metabolism, have been shown to either inhibit or activate the carboxylase. Inhibition by 6-phosphogluconate, NADPH, or fructose bisphosphate is pronounced when the enzyme is activated by preincubation at high levels of bicarbonate (20 to 50 mM) in the presence of magnesium (3,7,21). In contrast, upon incubation at low levels of bicarbonate (about 1 mM), the carboxylase is relatively inactive, and phosphorylated metabolites (particularly NADPH and phosphogluconate) markedly activate the inactive enzyme if included in the preincubation mixture (4-6, 8, 9, 15, 18). Such behavior yielded a plausible hypothesis for the physiological regulation of this key enzyme based upon the known concentrations and fluctuations of these metabolites within the chloroplast (1, 14).The cyanobacteria are procaryotic organisms that resemble the chloroplasts of photosynthetic eucaryotes in that they possess the capacity to split water and thus evolve oxygen via photosystem II-directed photosynthesis. Moreover, kinetic studies of "CO2 fixation and 32P incorporation into Aphanocapsa strains 6308 and 6714 (16, 17) showed a rapid increase in the concentration of phosphogluconate and changes in the concentrations of sugar mono-and diphosphates when cells wer...

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