Metabolic engineering of Escherichia coli for the biosynthesis of 2-pyrrolidone

• Published in: Metabolic engineering communications Vol. 3; no. C; pp. 1 - 7
• Main Authors: Zhang, Jingwei, Kao, Emily, Wang, George, Baidoo, Edward E.K., Chen, Matthew, Keasling, Jay. D.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.12.2016; Elsevier
• Subjects: 2-Pyrrolidone; 60 APPLIED LIFE SCIENCES; BASIC BIOLOGICAL SCIENCES; binding proteins; Biosynthesis; E. coli; Escherichia coli; genes; Glutamate; glutamate decarboxylase; glutamic acid; maltose; Metabolic engineering; mutants; polymers; solvents; Streptomyces; temperature; Metabolic engineering; Biosynthesis; Glutamate; 2-Pyrrolidone; E. coli
• ISSN: 2214-0301; 2214-0301
• DOI: 10.1016/j.meteno.2015.11.001

2-Pyrrolidone is a valuable bulk chemical with myriad applications as a solvent, polymer precursor and active pharmaceutical intermediate. A novel 2-pyrrolidone synthase, ORF27, from Streptomyces aizunensis was identified to catalyze the ring closing dehydration of γ-aminobutyrate. ORF27's tendency to aggregate was resolved by expression at low temperature and fusion to the maltose binding protein (MBP). Recombinant Escherichia coli was metabolically engineered for the production of 2-pyrrolidone from glutamate by expressing both the genes encoding GadB, a glutamate decarboxylase, and ORF27. Incorporation of a GadB mutant lacking H465 and T466, GadB_ΔHT, improved the efficiency of one-pot 2-pyrrolidone biosynthesis in vivo. When the recombinant E. coli strain expressing the E. coli GadB_ΔHT mutant and the ORF27-MBP fusion was cultured in ZYM-5052 medium containing 9g/L of l-glutamate, 7.7g/L of l-glutamate was converted to 1.1g/L of 2-pyrrolidone within 31h, achieving 25% molar yield from the consumed substrate. [Display omitted] •ORF27 from Streptomyces aizunensis catalyzes formation of 2-pyrrolidone from γ-aminobutyrate.•Recombinant Escherichia coli with GadB and ORF27 produces 2-pyrrolidone from glutamate.•Engineered strain capable of producing 1.1g/L of 2-pyrrolidone from 9g/L of glutamate within 31h.