Engineering cyanobacteria for increased growth and productivity

Increasing the photosynthetic efficiency is one of the strategies to increase the crop yields to meet the requirement of 50% more food by 2050. Due to the similarity on photosynthesis between crops and cyanobacteria, cyanobacteria are ideal alternatives to study photosynthesis since cyanobacteria are prokaryotes, easier to engineer and have shorter life cycle. On the other hand, cyanobacteria are promising cell factories for food additives, biofuels, and other products. To get the desired products from cyanobacteria directly will consume atmospheric CO2 and avoid additional releasing of CO2 from the usage of fossil resources. In this thesis, four CBB cycle enzymes were overexpressed individually in the model cyanobacterium Synechocystis PCC 6803. To get ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) overexpressed, two methods were used. One was to introduce another copy of the carboxysome protein CcmM gene into the cells since CcmM is essential for packing RuBisCO into the carboxysome. Another way was to tag the RuBisCO gene either on the N terminus of the large subunit or on the C terminus of the small subunit by FLAG. Even though the RuBisCO level increased, the specific RuBisCO activity did not change. Fructose-1,6-/sedoheptulose-1,7-bisphosphatase (FBP/SBPase), aldolase (FBA) and transketolase (TK) were overexpressed by introducing a second copy of corresponding gene. The engineered strains with increased levels of RuBisCO, FBP/SBPase, and FBA grew faster, had higher maximum net oxygen evolution rate and accumulated more biomass when cultivated under 100µmol photons m-2 s-1 light intensity. The strain carrying more TK showed a chlorotic phenotype but still accumulated more biomass under the same light condition. Four strains with one of the CBB cycle enzymes overexpressed were selected to investigate the effects on ethanol production. Increased ethanol production and ethanol to total biomass rate were observed in the CBB cycle engineered strains. The best strain produced almost 50% ethanol out of the total biomass. This work shows that overexpressing selected enzymes of the CBB cycle in cyanobacteria resulted in enhanced total biomass accumulation and increased compound (exampled as ethanol) production under certain growth conditions