Background: The disposal of plastic waste is a major environmental challenge. With recent advances in microbial genetic and metabolic engineering technologies, microbial polyhydroxyalkanoates (PHAs) are being used as next-generation biomaterials to replace petroleum-based synthetic plastics in a sustainable future. However, the relatively high production cost of bioprocesses hinders the production and application of microbial PHAs on an industrial scale.
Results: Here, we describe a rapid strategy to rewire metabolic networks in an industrial microorganism, Corynebacterium glutamicum, for the enhanced production of poly(3-hydroxybutyrate) (PHB). A three-gene PHB biosynthetic pathway in Rasltonia eutropha was refactored for high-level gene expression. A fluorescence-based quantification assay for cellular PHB content using BODIPY was devised for the rapid fluorescence-activated cell sorting (FACS)-based screening of a large combinatorial metabolic network library constructed in C. glutamicum. Rewiring metabolic networks across the central carbon metabolism enabled highly efficient production of PHB up to 29% of dry cell weight with the highest cellular PHB productivity ever reported in C. glutamicum using a sole carbon source.
Conclusions: We successfully constructed a heterologous PHB biosynthetic pathway and rapidly optimized metabolic networks across central metabolism in C. glutamicum for enhanced production of PHB using glucose or fructose as a sole carbon source in minimal media. We expect that this FACS-based metabolic rewiring framework will accelerate strain engineering processes for the production of diverse biochemicals and biopolymers.
Keywords: Combinatorial optimization; Corynebacterium glutamicum; Metabolic engineering; Poly(3-hydroxybutyrate).
© 2023. The Author(s).