Rapid combinatorial rewiring of metabolic networks for enhanced poly(3-hydroxybutyrate) production in Corynebacterium glutamicum

• Published in: Microbial cell factories Vol. 22; no. 1; p. 29
• Main Authors: Yim, Sung Sun, Choi, Jae Woong, Lee, Yong Jae, Jeong, Ki Jun
• Format: Journal Article
• Language: English
• Published: England BioMed Central Ltd 17.02.2023; BioMed Central; BMC
• Subjects: 3-Hydroxybutyric Acid; Bacteria; Biomaterials; Biomedical materials; Bioplastics; Biopolymers; Biosynthesis; Carbon; Carbon - metabolism; Carbon sources; Combinatorial analysis; Combinatorial optimization; Corynebacteria; Corynebacterium glutamicum; Corynebacterium glutamicum - genetics; Corynebacterium glutamicum - metabolism; Dry cells; Enzymes; Flow cytometry; Fluorescence; Gene expression; Libraries; Metabolic Engineering; Metabolic networks; Metabolic Networks and Pathways; Metabolism; Microorganisms; Networks; Physiological aspects; Plastic debris; Plastics; Poly-3-hydroxybutyrate; Polyhydroxyalkanoates; Polyhydroxyalkanoates - metabolism; Polyhydroxybutyrate; Production costs; Properties; Transcription factors; South Korea; Poly(3-hydroxybutyrate); Metabolic engineering; Combinatorial optimization; Corynebacterium glutamicum
• ISSN: 1475-2859; 1475-2859
• DOI: 10.1186/s12934-023-02037-x

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. 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. 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.