Introduction of a phenylalanine sink in fast growing cyanobacterium Synechococcus elongatus PCC 11801 leads to improved PSII efficiency, linear electron transport, and carbon fixation.

Cyanobacteria are investigated for fundamental photosynthesis research and sustainable production of valuable biochemicals. However, low product titer and biomass productivities are major bottlenecks to the economical scale-up. Recent studies have shown that the introduction of a metabolic sink, such as sucrose, 2,3-butanediol, and 2-phenyl ethanol, in cyanobacteria improves carbon fixation by relieving the "sink" limitation of photosynthesis. However, the impact of light intensity on the behavior of this sink-derived enhancement in carbon fixation is not well understood and is necessary for translation to outdoor cultivation. Here, using random mutagenesis, we engineered Synechococcus elongatus PCC 11801 to overproduce 1.24 g L phenylalanine (Phe) in 3 days, identified L531W in the TolC protein as an important driver of Phe efflux, and investigated the effect of light intensity on total carbon fixation. We found that low light results in competition between biomass and Phe, whereas under excess light, a higher flux of fixed carbon is directed to the Phe sink. The introduction of the Phe sink improves the quantum yields of photosystem I and II with a concomitant increase in the total electron flow leading to nearly 70% increase in carbon fixation at high light in the mutant strain. Additionally, the cyclic electron flow decreased, which has implications for the ATP/NADPH production ratio. Our data highlight how light intensity affects the sink-derived enhancement in carbon fixation, the role of CEF to balance the source-sink demand for ATP and NADPH, and the enhancement of inorganic carbon fixation in cyanobacteria with an engineered sink.