Abstract
The aim of this work was the establishment of a novel method to determine the metabolic load on host-cell metabolism resulting from recombinant protein production in Escherichia coli. This tool can be used to develop strategies to optimise recombinant fermentation processes through adjustment of recombinant-protein expression to the biosynthetic capacity of the host-cell. The signal molecule of the stringent-response network, guanosine tetraphosphate (ppGpp), and its precursor nucleotides were selected for the estimation of the metabolic load relating to recombinant-protein production. An improved analytical method for the quantification of nucleotides by ion-pair, high-performance liquid chromatography was established. The host-cell response upon overexpression of recombinant protein in fed-batch fermentations was investigated using the production of human superoxide dismutase (rhSOD) as a model system. E. coli strains with different recombinant systems (the T7 and pKK promoter system) exerting different loads on host-cell metabolism were analysed with regard to intracellular nucleotide concentration, rate of product formation and plasmid copy number.
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Received: 30 April 1999 / Received revision: 26 July 1999 / Accepted: 1 August 1999
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Cserjan-Puschmann, M., Kramer, W., Duerrschmid, E. et al. Metabolic approaches for the optimisation of recombinant fermentation processes. Appl Microbiol Biotechnol 53, 43–50 (1999). https://doi.org/10.1007/s002530051612
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DOI: https://doi.org/10.1007/s002530051612