Optimization of lysine metabolism in Corynebacterium glutamicum

Jakob Vang Rytter

    Research output: Book/ReportPh.D. thesis

    Abstract

    Commercial pig and poultry production use the essential amino acid lysine as a feed additive with the purpose of optimizing the feed utilization. Lysine is produced by a fermentation process involving either Corynebacterium glutamicum or Escherichia coli. The global annual production is around 1,000,000 tons. The aim of this project is to optimize the yield of lysine in C. glutamicum using metabolic engineering strategies.
    According to a genome scale model of C. glutamicum, theoretically there is much room for increasing the lysine yield (Kjeldsen and Nielsen 2009). Lysine synthesis requires NADPH, and increased NADPH availability is therefore a potential way to enhance lysine production. The generation of NADPH is mainly located in the pentose phosphate pathway (PPP). Using the genome scale model the phosphoglucoisomerase enzyme (PGI) has been identified as a possible bottleneck in the metabolism, which the project intends to eliminate. PGI catalyzes the conversion of alpha-D-glucose-6-phosphate to fructose-6-phosphate just downstream of the branch in the glycolysis, but it also catalyzes the reverse reaction. It is unknown whether up- or down-regulation of the pgi is required to increase the flux through the PPP, increasing the NADPH synthesis and enabling increased lysine production.
    Synthetic promoter libraries (SPL) enable fine tuning of the expression of genes. To test the feasibility of SPL in C. glutamicum four constitutive SPLs and one inducible SPL were constructed. The libraries were placed in front of a β-galactosidase gene and the activity of the enzyme was measured to quantify the different promoter strengths in the libraries. The results demonstrate that the SPL technology is an applicable technique for metabolic engineering in C. glutamicum.
    SPL was used to both up- and down-regulate the expression of pgi, and it was measured how the regulations affected the lysine production. In this thesis it is shown that under the applied growth conditions it is possible to increase the lysine yield by a factor of 1.6 from 0.0086 mole lysine ∙ mole glucose-1 to 0.0138 mole lysine ∙ mole glucose-1.
    Original languageEnglish
    Place of PublicationKgs. Lyngby
    PublisherTechnical University of Denmark
    Number of pages111
    Publication statusPublished - 2013

    Cite this

    Rytter, J. V. (2013). Optimization of lysine metabolism in Corynebacterium glutamicum. Technical University of Denmark.