The regulatory protein CP12 can bind glyceraldehyde 3-phosphate dehydrogenase (GapDH) and phosphoribulokinase (PRK) in oxygenic phototrophs, thereby switching on and off the flux through the Calvin-Benson cycle (CBC) under light and dark conditions, respectively. However, it can be assumed that CP12 is also regulating CBC flux under further conditions associated with redox changes. To prove this hypothesis, the mutant Δ of the model cyanobacterium sp. PCC 6803 was compared to wild type and different complementation strains. Fluorescence microscopy showed for the first time the kinetics of assembly and disassembly of the CP12-GapDH-PRK complex, which was absent in the mutant Δ. Metabolome analysis revealed d... More
The regulatory protein CP12 can bind glyceraldehyde 3-phosphate dehydrogenase (GapDH) and phosphoribulokinase (PRK) in oxygenic phototrophs, thereby switching on and off the flux through the Calvin-Benson cycle (CBC) under light and dark conditions, respectively. However, it can be assumed that CP12 is also regulating CBC flux under further conditions associated with redox changes. To prove this hypothesis, the mutant Δ of the model cyanobacterium sp. PCC 6803 was compared to wild type and different complementation strains. Fluorescence microscopy showed for the first time the kinetics of assembly and disassembly of the CP12-GapDH-PRK complex, which was absent in the mutant Δ. Metabolome analysis revealed differences in the contents of ribulose 1,5-bisphosphate and dihydroxyacetone phosphate, the products of the CP12-regulated enzymes GapDH and PRK, between wild type and mutant Δ under changing CO conditions. Growth of Δ was not affected at constant light under different inorganic carbon conditions, however, the addition of glucose inhibited growth in darkness as well as under diurnal conditions. The growth defect in the presence of glucose is associated with the inability of Δ to utilize external glucose. These phenotypes could be complemented by ectopic expression of the native CP12 protein, however, expression of CP12 variants with missing redox-sensitive cysteine pairs only partly restored the growth with glucose. These experiments indicated that the loss of GapDH-inhibition CP12 is more critical than PRK association. Measurements of the NAD(P)H oxidation revealed an impairment of light intensity-dependent redox state regulation in Δ Collectively, our results indicate that CP12-dependent regulation of the CBC is crucial for metabolic adjustment under conditions leading to redox changes such as diurnal conditions, glucose addition, and different CO conditions in cyanobacteria.
