Light-optimized growth of cyanobacterial cultures: Growth phases and productivity of biomass and secreted molecules in light-limited batch growth.

Cyanobacteria are photosynthetic microorganisms whose metabolism can be modified through genetic engineering for production of a wide variety of molecules directly from CO2, light, and nutrients. Diverse molecules have been produced in small quantities by engineered cyanobacteria to demonstrate the feasibility of photosynthetic biorefineries. Consequently, there is interest in engineering these microorganisms to increase titer and productivity to meet industrial metrics. Unfortunately, differing experimental conditions and cultivation techniques confound comparisons of strains and metabolic engineering strategies. In this work, we discuss the factors governing photoautotrophic growth and demonstrate nutritionally replete conditions in which a model cyanobacterium can be grown to stationary phase with light as the sole limiting substrate. We introduce a mathematical framework for understanding the dynamics of growth and product secretion in light-limited cyanobacterial cultures. Using this framework, we demonstrate how cyanobacterial growth in differing experimental systems can be easily scaled by the volumetric photon delivery rate using the model organisms Synechococcus sp. strain PCC7002 and Synechococcus elongatus strain UTEX2973. We use this framework to predict scaled up growth and product secretion in 1L photobioreactors of two strains of Synechococcus PCC7002 engineered for production of l-lactate or L-lysine. The analytical framework developed in this work serves as a guide for future metabolic engineering studies of cyanobacteria to allow better comparison of experiments performed in different experimental systems and to further investigate the dynamics of growth and product secretion.

Engineering photosynthetic production of L-lysine.

L-lysine and other amino acids are commonly produced through fermentation using strains of heterotrophic bacteria such as Corynebacterium glutamicum. Given the large amount of sugar this process consumes, direct photosynthetic production is intriguing alternative. In this study, we report the development of a cyanobacterium, Synechococcus sp. strain PCC 7002, capable of producing L-lysine with CO2 as the sole carbon-source. We found that heterologous expression of a lysine transporter was required to excrete lysine and avoid intracellular accumulation that correlated with poor fitness. Simultaneous expression of a feedback inhibition resistant aspartate kinase and lysine transporter were sufficient for high productivities, but this was also met with a decreased chlorophyll content and reduced growth rates. Increasing the reductant supply by using NH4+, a more reduced nitrogen source relative to NO3-, resulted in a two-fold increase in productivity directing 18% of fixed carbon to lysine. Given this advantage, we demonstrated lysine production from media formulated with a municipal wastewater treatment sidestream as a nutrient source for increased economic and environmental sustainability. Based on our results, we project that Synechococcus sp. strain PCC 7002 could produce lysine at areal productivities approaching that of sugar cane to lysine via fermentation using non-agricultural lands and low-cost feedstocks.