Effects of Overexpressing Calvin-Bassham-Benson Cycle Genes in Synechocystis PCC 6803
- Conference: Synthetic Biology Engineering Evolution Design SEED
- Year: 2017
- Proceeding: 2017 Synthetic Biology: Engineering, Evolution & Design (SEED)
- Group: Poster Session
Two different strategies were used to increase the RuBisCO content in the cells. One was to introduce ccmMgene into wild type cells on a self-replication vector (Liang and Lindblad, 2016) and the second was to add a FLAG tag sequence on the large subunit of the N terminus or the small subunit of the C terminus of RuBisCO. Since CcmM is a carboxysome protein that binds RuBisCO, CcmM may localize RuBisCO into the carboxysome as well as stabilize the enzyme. The presence of a FLAG tag enhanced transcription of the genes encoding RuBisCO, and, with high possibility, also enhanced the initiation of translation and/or stability of the enzyme. However, when using a streptavidin-binding tag II (strep-tag II), we did not observe similar effects. FBP/SBPase (di-functional enzyme in cyanobacteria), FBA and TK were successfully overexpressed by introducing another gene copy on self-replication vector (Liang and Lindblad, 2016).
The results demonstrate that overexpressing above discussed enzymes in Synechocystis resulted in increased in vivo oxygen evolution. Under 100Î¼mol m-2 s-1 irradiance, growth and biomass were enhanced in cells overexpressing RuBisCO, FBP/SBPase and FBA. Cells overexpressing TK behaved differently, showing slower growth but increased biomass accumulation. In addition, 20Î¼mol m-2 s-1 irradiance restricted the engineered strains' growth and biomass accumulation. However, cells overexpressing TK showed increased biomass under 20Î¼mol m-2 s-1 irradiance. This study demonstrates enhanced growth and photosynthesis in cells of the cyanobacterium Synechocystis overexpressing selected enzymes of the CBB cycle. RuBisCO, SBP/FBPase, FBA and TK are four potential targets to improve growth of, and yield of valuable products from, SynechocystisPCC 6803.
Feiyan Liang, Peter Lindblad*, Effects of Overexpressing Calvin-Bassham-Benson Cycle Carbon Flux Control Enzymes in the Cyanobacterium Synechocystis PCC 6803. (2016) Metabolic Engineering 38:56-64. doi: 10.1016/j.ymben.2016.06.005.