title: Engineering E. coli for the biosynthesis of 3-hydroxy-γ-butyrolactone (3HBL) and 3,4-dihydroxybutyric acid (3,4-DHBA) as value-added chemicals from glucose as a sole carbon source.
abstract: 3-hydroxy-γ-butyrolactone (3HBL) is a versatile chiral synthon, deemed a top value-added chemical from biomass by the DOE. We recently reported the first biosynthetic pathway towards 3HBL and its hydrolyzed form, 3,4-dihydroxybutyric acid (3,4-DHBA) in recombinant Escherichia coli using glucose and glycolic acid as feedstocks and briefly described their synthesis solely from glucose. Synthesis from glucose requires integration of the endogenous glyoxylate shunt with the 3,4-DHBA/3HBL pathway and co-overexpression of seven genes, posing challenges with respect to expression, repression of the glyoxylate shunt and optimal carbon distribution between the two pathways. Here we discuss engineering this integration. While appropriate media and over-expression of glyoxylate shunt enzymes helped overcome repression, two orthogonal expression systems were employed to address the expression and carbon distribution challenge. Synthesis of up to 0.3g/L of 3HBL and 0.7g/L of 3,4-DHBA solely from glucose was demonstrated, amounting to 24% of the theoretical maximum.
authors: Department of Chemical Engineering, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Room E17-504, Cambridge, MA 02139, USA; Synthetic Biology Engineering Research Center (SynBERC), Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors: Himanshu
authors: H
authors: Dhamankar
authors: Synthetic Biology Engineering Research Center (SynBERC), Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Microbiology Graduate Program, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors: Yekaterina
authors: Y
authors: Tarasova
authors: Department of Chemical Engineering, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Room E17-504, Cambridge, MA 02139, USA; Synthetic Biology Engineering Research Center (SynBERC), Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors: Collin H
authors: CH
authors: Martin
authors: Department of Chemical Engineering, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Room E17-504, Cambridge, MA 02139, USA; Synthetic Biology Engineering Research Center (SynBERC), Massachusetts Institute of Technology, Cambridge, MA 02139, USA. Electronic address: kljp@mit.edu.
authors: Kristala L J
authors: KL
authors: Prather
mesh_terms: D015107:4-Butyrolactone / Q000031:analogs & derivatives* / Q000096:biosynthesis; D002072:Butylene Glycols / Q000378:metabolism*; D002087:Butyrates / Q000378:metabolism*; D049109:Cell Proliferation / Q000502:physiology; D003198:Computer Simulation; D004926:Escherichia coli / Q000502:physiology*; D029968:Escherichia coli Proteins / Q000502:physiology*; D024861:Genetic Enhancement / Q000379:methods*; D005947:Glucose / Q000378:metabolism*; D060847:Metabolic Engineering / Q000379:methods*; D008954:Models, Biological
chemical_list: C014489:3,4-dihydroxybutyrate; C511346:3-hydroxy-gamma-butyrolactone; D002072:Butylene Glycols; D002087:Butyrates; D029968:Escherichia coli Proteins; D005947:Glucose; D015107:4-Butyrolactone
keywords: 3-hydroxy-γ-buytrolactone; Glycolate; Glyoxylate shunt; Hydroxyacids; Metabolic burden; Orthogonal expression systems; T7 RNAP based expression