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@PHDTHESIS{Milke:877282,
      author       = {Milke, Lars},
      title        = {{E}ngineering of {C}orynebacterium glutamicum towards
                      increased malonyl-{C}o{A} availability for polyketide
                      synthesis},
      volume       = {223},
      school       = {Heinrich-Heine-Universität Düsseldorf},
      type         = {Dr.},
      address      = {Jülich},
      publisher    = {Forschungszentrum Jülich GmbH Zentralbibliothek, Verlag},
      reportid     = {FZJ-2020-02105},
      isbn         = {978-3-95806-480-5},
      series       = {Schriften des Forschungszentrums Jülich. Reihe
                      Schlüsseltechnologien / Key Technologies},
      pages        = {IX, 117 S.},
      year         = {2020},
      note         = {Biotechnologie 1; Heinrich-Heine-Universität Düsseldorf,
                      Diss., 2020},
      abstract     = {Polyketides are a structurally highly diverse group of
                      natural products with interesting healthpromoting effects on
                      humans. Despite all structural differences, polyketides are
                      synthesized from simple CoA-activated carboxylic acid
                      derivatives, such as acetyl-CoA or malonyl-CoA following a
                      mechanism closely related to fatty acid biosynthesis.
                      Unfortunately, polyketides are only synthesized in small
                      quantities by the respective native organism. In contrast,
                      microbial polyketide synthesis using engineered bacteria is
                      a promising approach to get access to the desired products.
                      Against this background, $\textit{Corynebacterium
                      glutamicum}$ strains for the production of different plant
                      polyketides such as stilbenes and flavonoids have been
                      constructed recently. However, it soon became evident that
                      the intracellular availability of malonyl-CoA is limiting
                      the overall product formation in these strains. Hence, the
                      main goal of this thesis was to optimize the intracellular
                      malonyl-CoA availability in $\textit{C. glutamicum}$ by
                      metabolic engineering. Additionally, the tailored strains
                      should be used for establishing synthesis of
                      biotechnological interesting polyketides. The following
                      results were obtained: 1) Reduction of the citrate synthase
                      activity to 5.5 \% compared to the $\textit{C. glutamicum}$
                      wild type by exchanging the promotor of the encoding
                      $\textit{gltA}$ gene, reduced acetyl-CoA consumption via the
                      tricarboxylic acid cycle, which in turn improved malonyl-CoA
                      availability. Upon transcriptional deregulation of
                      $\textit{accBC}$ and $\textit{accD1}$ encoding the two
                      subunits of acetyl-CoA carboxylase, malonyl-CoA synthesis
                      from acetyl-CoA was drastically improved allowing for the
                      synthesis of 65 mg/L (0.24 mM) naringenin und 450 mg/L (1.97
                      mM) resveratrol. Furthermore, improving the glucose uptake
                      and elimination of anaplerotic pyruvate carboxylation
                      reaction further contributed to an improved intracellular
                      malonyl-CoA availability in the ultimately constructed
                      strain $\textit{C. glutamicum}$ M-CoA. 2) Through episomal
                      expression of genes encoding heterologous type III
                      polyketide synthases from various plant species in the
                      constructed strain $\textit{C. glutamicum}$ M-CoA, microbial
                      synthesis of a pentaketide (noreugenin) but also
                      phenylbutanoids (raspberry ketone, zingerone, benzylacetone)
                      with a $\textit{ldhA}$-deficient variant could be
                      established. The respective strains allowed for the
                      synthesis of up to 53.3 mg/L (0.28 mM) noreugenin, 100 mg/L
                      (0.61 mM) raspberry ketone, 70 mg/L (0.36 mM) zingerone and
                      10.5 mg/L (0.07 mM) benzylacetone from simple precursor
                      molecules, respectively. 3) Hitherto, only type III
                      polyketides can be synthesized by engineered $\textit{C.
                      glutamicum}$ strains. In the context of this study,
                      functional expression of a codon-optimized gene variant
                      encoding the type I polyketide synthase 6-methylsalicylic
                      acid synthase ChlB1 from $\textit {Streptomyces
                      antibioticus}$ of 1,756 amino acids size was achieved. This
                      allowed for the synthesis of up to 41 mg/L (0.27 mM)
                      6-methylsalicylic acid. It was found that $\textit{C.
                      glutamicum}$ has an endogenous
                      phosphopantetheinyltransferase activity, which can
                      post-translationally activate ChlB1. This makes $\textit{C.
                      glutamicum}$ a promising host for the production of other
                      interesting type I polyketides.},
      cin          = {IBG-1},
      cid          = {I:(DE-Juel1)IBG-1-20101118},
      pnm          = {581 - Biotechnology (POF3-581)},
      pid          = {G:(DE-HGF)POF3-581},
      typ          = {PUB:(DE-HGF)3 / PUB:(DE-HGF)11},
      urn          = {urn:nbn:de:0001-2020072314},
      url          = {https://juser.fz-juelich.de/record/877282},
}