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Biosynthesis of polyhydroxyalkanoates containing 2-hydroxybutyrate from unrelated carbon source by metabolically engineered Escherichia coli

  • Applied genetics and molecular biotechnology
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Abstract

We have previously reported in vivo biosynthesis of polylactic acid (PLA) and poly(3-hydroxybutyrate-co-lactate) [P(3HB-co-LA)] employing metabolically engineered Escherichia coli strains by the introduction of evolved Clostridium propionicum propionyl-CoA transferase (Pct Cp ) and Pseudomonas sp. MBEL 6-19 polyhydroxyalkanoate (PHA) synthase 1 (PhaC1 Ps6-19). Using this in vivo PLA biosynthesis system, we presently report the biosynthesis of PHAs containing 2-hydroxybutyrate (2HB) monomer by direct fermentation of a metabolically engineered E. coli strain. The recombinant E. coli ldhA mutant XLdh strain expressing PhaC1 Ps6-19 and Pct Cp was developed and cultured in a chemically defined medium containing 20 g/L of glucose and varying concentrations of 2HB and 3HB. PHAs consisting of 2HB, 3HB, and a small fraction of lactate were synthesized. Their monomer compositions were dependent on the concentrations of 2HB and 3HB added to the culture medium. Even though the ldhA gene was completely deleted in the chromosome of E. coli, up to 6 mol% of lactate was found to be incorporated into the polymer depending on the culture condition. In order to synthesize PHAs containing 2HB monomer without feeding 2HB into the culture medium, a heterologous metabolic pathway for the generation of 2HB from glucose was constructed via the citramalate pathway, in which 2-ketobutyrate is synthesized directly from pyruvate and acetyl-CoA. Introduction of the Lactococcus lactis subsp. lactis Il1403 2HB dehydrogenase gene (panE) into E. coli allowed in vivo conversion of 2-ketobutyrate to 2HB. The metabolically engineered E. coli XLdh strain expressing the phaC1437, pct540, cimA3.7, and leuBCD genes together with the L. lactis Il1403 panE gene successfully produced PHAs consisting of 2HB, 3HB, and a small fraction of lactate by varying the 3HB concentration in the culture medium. As the 3HB concentration in the medium increased the 3HB monomer fraction in the polymer, the polymer content increased. When Ralstonia eutropha phaAB genes were additionally expressed in this recombinant E. coli XLdh strain, P(2HB-co-3HB-co-LA) having small amounts of 2HB and LA monomers could also be produced from glucose as a sole carbon source. The metabolic engineering strategy reported here should be useful for the production of PHAs containing 2HB monomer.

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Acknowledgements

This work was supported by the Korean Systems Biology Research Project (20090065571) of the Ministry of Education, Science and Technology (MEST) through the National Research Foundation (NRF) of Korea. Further support by World Class University program (R32-2008-000-10142-0) and by the Advanced Biomass R&D Center of Korea (ABC-2010-0029799) through the Global Frontier Research Program of MEST is appreciated. B.K.S, S.H.L, and S.J.P appreciate the financial supports from the R&D Program of MKE/KEIT (10032001) and KRICT.

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Authors and Affiliations

  1. Chemical Biotechnology Research Center, Korea Research Institute of Chemical Technology, P.O. Box 107, Sinseongno 19, Yuseong-gu, Daejeon, 305-600, Republic of Korea

    Si Jae Park, Seung Hwan Lee & Bong Keun Song

  2. Metabolic and Biomolecular Engineering National Research Laboratory, Department of Chemical and Biomolecular Engineering (BK21 Program), Center for Systems and Synthetic Biotechnology, and Institute for the BioCentury, KAIST, 291 Daehak-ro, Yuseong-gu, Daejeon, 305-701, Republic of Korea

    Tae Woo Lee, Min Kyung Kim & Sang Yup Lee

  3. Department of Bio and Brain Engineering, Department of Biological Sciences, BioProcess Engineering Research Center and Bioinformatics Research Center, KAIST, 291 Daehak-ro, Yuseong-gu, Daejeon, 305-701, Republic of Korea

    Sang Yup Lee

  4. Corporate R&D, LG Chem Research Park, 104-1 Moonji-dong, Yuseong-gu, Daejeon, 305-380, Republic of Korea

    Sung-Chul Lim

  5. Marine Biotechnology Research Center, Korea Ocean Research & Development Institute, PO Box 29, Ansan, 425-600, Republic of Korea

    Tae Wan Kim

  6. Medicinal Chemistry Research Center, Korea Research Institute of Chemical Technology, P.O.Box 107, Sinseongno 19, Yuseong-gu, Daejeon, 305-600, Republic of Korea

    Hyuk Lee

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  1. Si Jae Park
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Correspondence to Sang Yup Lee.

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Park, S.J., Lee, T.W., Lim, SC. et al. Biosynthesis of polyhydroxyalkanoates containing 2-hydroxybutyrate from unrelated carbon source by metabolically engineered Escherichia coli . Appl Microbiol Biotechnol 93, 273–283 (2012). https://doi.org/10.1007/s00253-011-3530-x

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  • DOI: https://doi.org/10.1007/s00253-011-3530-x

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