The Adc1 Knockout with ProC Overexpression in Synechocystis Sp. PCC 6803 Induces a Diversion of Acetyl-CoA to Produce More Polyhydroxybutyrate

Background: Lack of nutrients, in particular nitrogen and phosphorus, has been known in the field to sense glutamate production via 2-oxoglutarate and subsequently accelerate carbon storage, including glycogen and polyhydroxybutyrate (PHB), in cyanobacteria, but a few studies have focused on arginine catabolism. In this study, we first time demonstrated that gene manipulation on proC and adc1, related to proline and polyamine syntheses in arginine catabolism, had a significant impact on enhanced PHB production during late growth phase and nutrient-modified conditions. We constructed Synechocystis sp. PCC 6803 with an overexpressing proC gene, encoding Δpyrroline-5-carboxylate reductase in proline production, and adc1 disruption resulted in lower polyamine synthesis.

Results: Three engineered Synechocystis sp. PCC 6803 strains, including a ProC-overexpressing strain (OXP), adc1 mutant, and an OXP strain lacking the adc1 gene (OXP/Δadc1), certainly increased the PHB accumulation under nitrogen and phosphorus deficiency. The possible advantages of single proC overexpression include improved PHB and glycogen storage in late phase of growth and long-term stress situations. However, on day 7 of treatment, the synergistic impact created by OXP/Δadc1 increased PHB synthesis by approximately 48.9% of dry cell weight, resulting in a shorter response to nutrient stress than the OXP strain. Notably, changes in proline and glutamate contents in engineered strains, in particular OXP and OXP/Δadc1, not only partially balanced the intracellular C/N metabolism but also helped cells acclimate under nitrogen (N) and phosphorus (P) stress with higher chlorophyll a content in comparison with wild-type control.

Conclusions: In Synechocystis sp. PCC 6803, overexpression of proC resulted in a striking signal to PHB and glycogen accumulation after prolonged nutrient deprivation. When combined with the adc1 disruption, there was a notable increase in PHB production, particularly in situations where there was a strong C supply and a lack of N and P.