On-line estimation of the metabolic burden resulting from the synthesis of plasmid-encoded and heat-shock proteins by monitoring respiratory energy generation

Authors

  • Frank Hoffmann,

    1. Biochemical Engineering Division, GBF German Research Center for Biotechnology, Mascheroder Weg 1, 38124 Braunschweig, Germany; telephone: 0049-531-6181; fax: 0049-531-6181-111
    Search for more papers by this author
    • Present address of F. Hoffmann: Institut für Biotechnologie, Martin-Luther-Universität Halle-Wittenberg, Kurt-Mothers-Str. 3, 06120 Halle/Saale, Germany

  • Ursula Rinas

    Corresponding author
    1. Biochemical Engineering Division, GBF German Research Center for Biotechnology, Mascheroder Weg 1, 38124 Braunschweig, Germany; telephone: 0049-531-6181; fax: 0049-531-6181-111
    • Biochemical Engineering Division, GBF German Research Center for Biotechnology, Mascheroder Weg 1, 38124 Braunschweig, Germany; telephone: 0049-531-6181; fax: 0049-531-6181-111
    Search for more papers by this author

  • This article is dedicated to James Bailey, whose pioneering work stimulated the research presented in this article.

Abstract

Human basic fibroblast growth factor (hFGF-2) was produced in high-cell density cultures of recombinant Escherichia coli using a temperature-inducible expression system. The synthesis rates of proteins were followed by two-dimensional gel electrophoresis of the 35S-methionine-labeled proteom. After temperature induction of hFGF-2 synthesis, the rate of total protein synthesis per biomass increased by a factor of three, mainly as a result of the additional synthesis of hFGF-2 and heat-shock proteins. The synthesis rates of heat-shock proteins and constitutive plasmid-encoded proteins increased after the temperature upshift also in the control strain without hFGF-2 gene but followed time profiles different from the producing strain. The energy demand for the extra synthesis of plasmid-encoded and heat-shock proteins resulted in an elevated respiratory activity and, consequently, in a reduction of the growth rate and the biomass yield. A procedure was developed to relate the energy demand for the additional synthesis of these proteins to the generation of energy in the respiratory pathway. Specific energy production was estimated based on on-line measurable rates of oxygen consumption, or carbondioxide evolution and growth, respectively. In this way, the metabolic burden resulting from the synthesis of plasmid-encoded and heat-shock proteins was quantified from on-line accessible data. © 2001 John Wiley & Sons, Inc. Biotechnol Bioeng 76: 333–340, 2001.

Ancillary