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Development of sucrose-utilizing Escherichia coli K-12 strain by cloning β-fructofuranosidases and its application for l-threonine production

Applied Microbiology and Biotechnology volume 88, pages 905–913 (2010)Cite this article

Abstract

Sucrose is one of the most promising carbon sources for industrial fermentation. To achieve sucrose catabolism, the sucrose utilization operons have been introduced into microorganisms that are not able to utilize sucrose. However, the rates of growth and sucrose uptake of these engineered strains were relatively low to be successfully employed for industrial applications. Here, we report a practical example of developing sucrose-utilizing microorganisms using Escherichia coli K-12 as a model system. The sucrose utilizing ability was acquired by introducing only β-fructofuranosidase from three different sucrose-utilizing organisms (Mannheimia succiniciproducens, E. coli W, and Bacillus subtilis). Among them, the M. succiniciproducens β-fructofuranosidase was found to be the most effective for sucrose utilization. Analyses of the underlying mechanism revealed that sucrose was hydrolyzed into glucose and fructose in the extracellular space and both liberated hexoses could be transported by their respective uptake systems in E. coli K-12. To prove that this system can also be applied for the production of useful metabolites, the M. succiniciproducens β-fructofuranosidase was introduced into the engineered l-threonine production strain of E. coli K-12. This recombinant strain was able to produce 51.1 g/L l-threonine by fed-batch culture, resulting in an overall yield of 0.284 g l-threonine per g sucrose. This simple approach to make E. coli K-12 to acquire sucrose-utilizing ability and its successful biotechnological application can be employed to develop sustainable bioprocesses using renewable biomass.

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Acknowledgements

This work was supported by the Korea–Australia Collaborative Research Project on the Development of Sucrose-Based Bioprocess Platform (10030795) from the Korean Ministry of Knowledge Economy. Further support by World Class University program (R322009000101420) by the Ministry of Education, Science and Technology through the National Research Foundation is appreciated.

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Authors and Affiliations

  1. Metabolic and Biomolecular Engineering National Research Laboratory, Department of Chemical and Biomolecular Engineering (BK21 Program), BioProcess Engineering Research Center, Center for Systems and Synthetic Biotechnology, and Institute for the BioCentury, KAIST, Daejeon, 305-701, Republic of Korea

    Jeong Wook Lee, Sol Choi, Jin Hwan Park & Sang Yup Lee

  2. Department of Bio and Brain Engineering and Bioinformatics Research Center, Department of Biological Sciences, KAIST, Daejeon, 305-701, Republic of Korea

    Sang Yup Lee

  3. Australian Institute for Bioengineering and Nanotechnology (AIBN), The University of Queensland, St. Lucia, QLD, 4072, Australia

    Claudia E. Vickers & Lars K. Nielsen

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  1. Jeong Wook Lee
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  2. Sol Choi
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  3. Jin Hwan Park
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  4. Claudia E. Vickers
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  5. Lars K. Nielsen
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  6. Sang Yup Lee
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Correspondence to Jin Hwan Park or Sang Yup Lee.

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Lee, J.W., Choi, S., Park, J.H. et al. Development of sucrose-utilizing Escherichia coli K-12 strain by cloning β-fructofuranosidases and its application for l-threonine production. Appl Microbiol Biotechnol 88, 905–913 (2010). https://doi.org/10.1007/s00253-010-2825-7

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  • DOI: https://doi.org/10.1007/s00253-010-2825-7

Keywords

  • Sucrose
  • β-Fructofuranosidase
  • Sucrose 6-phosphate hydrolase
  • Sucrase
  • l-threonine