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@PHDTHESIS{Buchholz:33319,
      author       = {Buchholz, Arne Karsten},
      title        = {{Q}uantifizierung intrazellulärer {M}etabolitdynamiken zur
                      {U}ntersuchung mikrobieller {S}toffwechselnetzwerke},
      volume       = {3971},
      issn         = {0944-2952},
      school       = {Univ. Bonn},
      type         = {Dr. (Univ.)},
      address      = {Jülich},
      publisher    = {Forschungszentrum Jülich GmbH Zentralbibliothek, Verlag},
      reportid     = {PreJuSER-33319, Juel-3971},
      series       = {Berichte des Forschungszentrums Jülich},
      pages        = {IV, 198 p.},
      year         = {2002},
      note         = {Record converted from VDB: 12.11.2012; Bonn, Univ., Diss.,
                      2001},
      abstract     = {The rational improvement of the productivity of cellular
                      systems is one of the main goals of modern biotechnology and
                      may be accomplished in two ways. One approach is the
                      optimization of the production process ("bioprocess
                      engineering"). Another possibility is the improvement of the
                      cellular system itself by manipulatiog the metabolic
                      pathways ("metabolic engineering") . A precise manipulation
                      of microbial metabolism, and thereby a rational strain
                      development, is possible today by using the tools of
                      molecular biology. However, a prerequisite of this
                      manipulation is a detailed and quantitative knowledge of the
                      dynamics and regulation of metabolic fluxes in microbial
                      metabolism. In this work a dynamic method is used for the
                      quantification of microbial metabolism. The microorganism is
                      cultivated in a substrate limited steady state and then
                      rapidly shifted away from this metabolic equilibrium by
                      application of a substrate pulse ("pulse experiment").
                      Samples are taken using a rapid sampling and quenching
                      technique, which was developed in earlier works. The
                      metabolites are then extracted and analyzed to determine the
                      metabolic response ofthe microorganism. Using a novel system
                      for injecting substrate pulses, a number of dynamic pulse
                      experiments were performed. Intracellular metabolite
                      dynamics were quantified using newly developed analytical
                      techniques based an HPLC-MS, UV-HPLC, CE and enzymatic
                      assays. . Pulse experiments were carried out to investigate
                      the glucose uptake and utilisation via the PEP:PTS in
                      $\textit{Escherichia coli}$ K12. The scope of these
                      investigations was expanded using a genetically modified,
                      PTS-deficient strain $\textit{E. coli}$ 3pMK7 in order to
                      determine the metabolic changes taking place when the
                      Substrate uptake depends an ATP instead of PEP.
                      Additionally, the utilisation of glycerol by $\textit{E.
                      coli}$ K12 and the dependency of glycerol uptake an ATP was
                      investigated. In a further experiment, the switch from
                      glycerol to glucose as carbon source during a pulse
                      experiment showed a strong limitation of the available PEP
                      concentration during uptake. The time courses of
                      intracellular metabolite concentrations from these pulse
                      experiments are the basis for the development of kinetic
                      (structured) metabolic models. The modelling of the
                      experimental data allowed a quantification of metabolic
                      phenomena which up until now could only be explained
                      phenomenologically.},
      cin          = {IBT-2},
      cid          = {I:(DE-Juel1)VDB56},
      pnm          = {Verfahrenstechnik zur mikrobiellen Gewinnung von
                      Primärmetaboliten},
      pid          = {G:(DE-Juel1)FUEK93},
      typ          = {PUB:(DE-HGF)11 / PUB:(DE-HGF)3},
      url          = {https://juser.fz-juelich.de/record/33319},
}