Disrupted biosynthesis of glycogen and PHB combined with physicochemical factors increased bioproduction of γ-aminobutyric acid and δ-aminolevulinic acid in cyanobacteria

In Synechocystis sp. PCC 6803, redirecting the metabolic flow from upstream substrate for carbon metabolism i.e. glyceraldehyde-3-phosphate (the glycogen synthesis precursor) and poly(3-hydroxybutyrate) (PHB), towards ALA or GABA has not been studied. The PHB synthesis, one of the pathways consuming carbon via glycolysis pathway, was disrupted to generate the strain, ΔP, and another adjacent carbon utilizing pathway that is responsible for glycogen synthesis was also disrupted in the earlier mutant strain to generate ΔGP with the aim to increase carbon flux towards precursor metabolites for GABA and ALA biosynthesis. The ΔP strain showed an increase in intracellular glutamate and ALA levels (~19.5 and 2.5 ng g−1 DCW, respectively), whereas intracellular GABA levels were found increased (up to 7 ng g−1 DCW) in ΔGP strain as compared to the wild type strain (WT). Modified nutrient conditions including glucose, glutamate and LA supplementation as well as N-deprivation led to several fold increase in GABA and ALA levels in ΔP and ΔGP strains in comparison to those observed in WT, with the maximum GABA and ALA accumulation (~262 and 48 ng g−1 DCW, respectively) noticed in ΔP. Abiotic stresses including cold and UV irradiation also significantly improved the intracellular GABA and ALA accumulation in mutant strains. Increased biosynthesis of GABA and ALA in mutant strains was found correlated with the increased mRNA levels of genes involved in GABA, ALA and TCA cycle metabolism. Further organic acids profiling revealed that the disruption of two pathways redirects carbon routes to TCA cycle without affecting the viability of cells. This strategy might be applicable in strain development for enhancing other high value bioproducts synthesized by connected metabolic pathways that utilize carbon sources diverted from non-essential metabolism.