Metabolic engineering of Escherichia coli for biosynthesis of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) from glucose

Jung Eun Yang, Yong Jun Choi, Se Jin Lee, Kyoung Hee Kang, Hyuk Lee, Young Hoon Oh, Seung Hwan Lee, Si Jae Park, Sang Yup Lee

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69 Scopus citations

Abstract

The Escherichia coli XL1-blue strain was metabolically engineered to synthesize poly(3-hydroxybutyrate-co-3-hydroxyvalerate) [P(3HB-co-3HV)] through 2-ketobutyrate, which is generated via citramalate pathway, as a precursor for propionyl-CoA. Two different metabolic pathways were examined for the synthesis of propionyl-CoA from 2-ketobutyrate. The first pathway is composed of the Dickeya dadantii 3937 2-ketobutyrate oxidase or the E. coli pyruvate oxidase mutant (PoxB L253F V380A) for the conversion of 2-ketobutyrate into propionate and the Ralstonia eutropha propionyl-CoA synthetase (PrpE) or the E. coli acetyl-CoA:acetoacetyl-CoA transferase for further conversion of propionate into propionyl-CoA. The second pathway employs pyruvate formate lyase encoded by the E. coli tdcE gene or the Clostridium difficile pflB gene for the direct conversion of 2-ketobutyrate into propionyl-CoA. As the direct conversion of 2-ketobutyrate into propionyl-CoA could not support the efficient production of P(3HB-co-3HV) from glucose, the first metabolic pathway was further examined. When the recombinant E. coli XL1-blue strain equipped with citramalate pathway expressing the E. coli poxB L253F V380A gene and R. eutropha prpE gene together with the R. eutropha PHA biosynthesis genes was cultured in a chemically defined medium containing 20 g/L of glucose as a sole carbon source, P(3HB-co-2.3 mol% 3HV) was produced up to the polymer content of 61.7 wt.%. Moreover, the 3HV monomer fraction in P(3HB-co-3HV) could be increased up to 5.5 mol% by additional deletion of the prpC and scpC genes, which are responsible for the metabolism of propionyl-CoA in host strains.

Original languageEnglish
Pages (from-to)95-104
Number of pages10
JournalApplied Microbiology and Biotechnology
Volume98
Issue number1
DOIs
StatePublished - Jan 2014

Bibliographical note

Funding Information:
Acknowledgments This work was supported by the Technology Development Program to Solve Climate Changes (Systems Metabolic Engineering for Biorefineries) from the Ministry of Science, ICT, and Future Planning through the National Research Foundation of Korea (NRF-2012-C1AAA001-2012M1A2A2026556).

Keywords

  • 2-ketobutyrate
  • 2-ketobutyrate oxidase
  • Poly(3-hydroxybutyrate-co-3-hydroxyvalerate)
  • Polyhydroxyalkanoate
  • Propionyl-CoA
  • Propionyl-CoA synthetase

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