A Post-translational Metabolic Switch Enables Complete Decoupling of Bacterial Growth from Biopolymer Production in Engineered Escherichia coli

ACS Synth Biol. 2018 Nov 16;7(11):2686-2697. doi: 10.1021/acssynbio.8b00345. Epub 2018 Oct 22.


Most of the current methods for controlling the formation rate of a key protein or enzyme in cell factories rely on the manipulation of target genes within the pathway. In this article, we present a novel synthetic system for post-translational regulation of protein levels, FENIX, which provides both independent control of the steady-state protein level and inducible accumulation of target proteins. The FENIX device is based on the constitutive, proteasome-dependent degradation of the target polypeptide by tagging with a short synthetic, hybrid NIa/SsrA amino acid sequence in the C-terminal domain. Protein production is triggered via addition of an orthogonal inducer ( i.e., 3-methylbenzoate) to the culture medium. The system was benchmarked in Escherichia coli by tagging two fluorescent proteins (GFP and mCherry), and further exploited to completely uncouple poly(3-hydroxybutyrate) (PHB) accumulation from bacterial growth. By tagging PhaA (3-ketoacyl-CoA thiolase, first step of the route), a dynamic metabolic switch at the acetyl-coenzyme A node was established in such a way that this metabolic precursor could be effectively redirected into PHB formation upon activation of the system. The engineered E. coli strain reached a very high specific rate of PHB accumulation (0.4 h-1) with a polymer content of ca. 72% (w/w) in glucose cultures in a growth-independent mode. Thus, FENIX enables dynamic control of metabolic fluxes in bacterial cell factories by establishing post-translational synthetic switches in the pathway of interest.

Keywords: Escherichia coli; PHB; metabolic engineering; pathway engineering; proteolysis; synthetic biology.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acetyl Coenzyme A / genetics
  • Acetyl Coenzyme A / metabolism
  • Coenzyme A-Transferases / genetics
  • Coenzyme A-Transferases / metabolism
  • Escherichia coli / genetics
  • Escherichia coli / growth & development
  • Escherichia coli / metabolism*
  • Escherichia coli Proteins / metabolism
  • Hydroxybutyrates / metabolism*
  • Metabolic Engineering / methods*
  • Polyesters / metabolism*
  • Protein Processing, Post-Translational
  • Proteolysis


  • Escherichia coli Proteins
  • Hydroxybutyrates
  • Polyesters
  • poly-beta-hydroxybutyrate
  • Acetyl Coenzyme A
  • Coenzyme A-Transferases
  • 3-ketoacid CoA-transferase