Further enhanced acetate production by overexpressing selected CBB-cycle enzymes in an engineered Synechocystis PCC 6803 strain

Cyanobacteria are promising microorganisms as cell factories due to their ability to perform oxygenic photosynthesis. During this process the cell produces metabolites through CO 2 fixation powered by light energy. The main carbon fixation pathway in cyanobacteria is the Calvin-Benson-Bassham (CBB) cycle. Cyanobacteria, for example Synechocystis PCC 6803 (thereafter Synechocystis ) have the ability to produce carbon compounds, for example acetate, as side products of their metabolism. Acetate production can be increased through the insertion of a phosphoketolase and overexpression of a phosphotransacetylase. In the present study the production of acetate was further tuned by overexpression of selected enzyme(s) of CBB-cycle. The enzymes selected was aldolase (FBA) and its combination with either fructose-1,6/sedoheptulose-1,7-bisphosphatase (FBP/SBPase) or transketolase (TK). The higher increase was noticed in the strain overexpressing FBA, 1.5 times fold increase, followed by the strain overexpressing both FBA and FBP/SBPase, while the overexpression of both FBA and TK did not influence the acetate production. However, when CP12, a small regulatory protein of the CBB-cycle, was knocked out, the strain overexpressing FBA and TK showed increased acetate titers while the other two combinations showed moderate increase. These results indicate the capability to optimize the acetate production through overexpression of FBA and emphasize the dynamic regulation of the CBB-cycle enzyme(s).