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Production of polyhydroxyalkanoates by Escherichia coli mutants with defected mixed acid fermentation pathways

  • Applied Microbial and Cell Physiology
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Abstract

A series of Escherichia coli BW25113 mutants with reduced mixed acid fermentation were constructed. Genes ackA-pta, poxB, ldhA, adhE, and pflB encoding acetate kinase, phosphate acetyltransferase, pyruvate oxidase, d-lactate dehydrogenase, acetaldehyde dehydrogenase, and pyruvate formate-lyase, respectively, were deleted successively. When grown under microaerobic condition, the mutants reduced approximately 90% acetate excretion after the deletion of genes ackA-pta and poxB. Production of lactate, ethanol, and formate was also significantly reduced after the deletion of genes ldhA, adhE, and pflB, respectively. The accumulation of biomass and poly(3-hydroxybutyrate) (PHB) were significantly enhanced after deleting the mixed acid fermentation. E. coli mutant BWapld with deletions of ackA-pta, poxB, ldhA, and adhE produced twice the cell dry weight (CDW) and 3.5 times of PHB compared with its wild-type under microaerobic conditions. E. coli mutant BWapl with deletions of ackA-pta, poxB, and ldhA also achieved nearly twice CDW and three times of PHB content in comparison to the wild-type during 48 h static cultivation. Production of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) [P(3HB-co-3HV)] was observed in the mutants under static cultivation. E. coli mutant BWapld could produce approximately 50 wt.% P(3HB-co-3HV) consisting of 5 mol% of 3-hydroxyvalerate (3HV) under aerobic conditions, when the seed culture was inoculated at an appropriate time. When ackA-pta, poxB, ldhA, adhE, and pflB were deleted, E. coli mutant BWapldf accumulated over 70 wt.% P(3HB-co-3HV) consisting of 8 mol% 3HV under aerobic conditions.

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Acknowledgements

We appreciate the kind donation of strain E. coli BW25113 and plasmids pKD13, pKD46, and pCP20 from Dr. BL Wanner and the Coli Genetic Stock Center (CGSC), Yale University. Plasmid pBHR68 was generously provided by Professor Alexander Steinbüchel of the University of Münster in Germany. This research was financially supported by National High Tech 863 Grants (Project No. 20071860338), the State Basic Science Foundation 973 (Grant No. 2007CB707804), National Key Technology R&D Program (Project No. 20091851262), and Tsinghua University Initiative Scientific Research Program 2009THZ01005.

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Authors and Affiliations

  1. Protein Science Laboratory of the Ministry of Education, Department of Biology, School of Life Sciences, Tsinghua University, Beijing, 100084, China

    Jia Jian, Shao-Qin Zhang, Guo-Qiang Chen & Qiong Wu

  2. Department of Chemistry, University of Melbourne, Parkville, Melbourne, VIC 3052, Australia

    Zhen-Yu Shi

  3. Life Science Division, Graduate School at Shenzhen, Tsinghua University, Shenzhen, 518055, China

    Wei Wang

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  1. Jia Jian
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Correspondence to Guo-Qiang Chen or Qiong Wu.

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Jian, J., Zhang, SQ., Shi, ZY. et al. Production of polyhydroxyalkanoates by Escherichia coli mutants with defected mixed acid fermentation pathways. Appl Microbiol Biotechnol 87, 2247–2256 (2010). https://doi.org/10.1007/s00253-010-2706-0

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