Introduction of an alternative carbon fixation cycle into Synechocystis sp. PCC 6803

The aim of thesis was to replace the carbon-fixing Calvin cycle of Synechocystis sp. PCC 6803 with the 3-hydroxypropionate bi-cycle. A genome-scale metabolic model was made to test how integration of the 3-hydroxypropionate bi-cycle into the cyanobacterial cell might affect growth and central metabolism. Two separate approaches for knocking out the Calvin cycle were tested, by deleting either the phosphoribulokinase gene or replacing the RuBisCO operon with a metabolic bypass. The first DNA constructs for expression of all enzymes in the bi-cycle were made and introduced into Synechocystis sp. PCC 6803, although expression of proteins other than the malonyl-CoA reductase and propionyl-CoA synthase was not detected. Several enzymes of the bi-cycle were cloned and expressed in Escherichia coli for characterisation and protein crystallisation studies. The structure of the native state of the tri-functional malyl-CoA lyase involved in the 3-hydroxypropionate bi-cycle was solved. The efforts made in these studies should prove useful for the successful engineering of the remainder of the 3-hydroxypropionate bi-cycle into Synechocystis sp. PCC 6803.Open Acces