Metabolic adaptation of Escherichia coli during temperature-induced recombinant protein production: 2. Redirection of metabolic fluxes
Article first published online: 4 SEP 2002
Copyright © 2002 Wiley Periodicals, Inc.
Biotechnology and Bioengineering
Volume 80, Issue 3, pages 320–330, 5 November 2002
How to Cite
Weber, J., Hoffmann, F. and Rinas, U. (2002), Metabolic adaptation of Escherichia coli during temperature-induced recombinant protein production: 2. Redirection of metabolic fluxes. Biotechnol. Bioeng., 80: 320–330. doi: 10.1002/bit.10380
- Issue published online: 4 SEP 2002
- Article first published online: 4 SEP 2002
- Manuscript Accepted: 18 APR 2002
- Manuscript Received: 12 DEC 2001
- recombinant protein;
- metabolic burden;
- Escherichia coli;
- metabolic flux analysis
The impact of temperature-induced synthesis of human basic fibroblast growth factor (hFGF-2) in high-cell-density cultures of recombinant Escherichia coli was studied by estimating metabolic flux variations. Metabolic flux distributions in E. coli were calculated by means of a stoichiometric network and linear programming. After the temperature upshift, a substantially elevated energy demand for synthesis of hFGF-2 and heat shock proteins resulted in a redirection of metabolic fluxes. Catabolic pathways like the Embden-Meyerhof-Parnas pathway and the tricarboxylic acid (TCA) cycle showed significantly enhanced activities, leading to reduced flux to growth-associated pathways like the pentose phosphate pathway and other anabolic pathways. Upon temperature upshift, an excess of NADPH was produced in the TCA cycle by isocitrate dehydrogenase. The metabolic model predicted the involvement of a transhydrogenase generating additional NADH from NADPH, thereby increasing ATP regeneration in the respiratory chain. The influence of the temperature upshift on the host's metabolism was investigated by means of a control strain harboring the “empty” parental expression vector. The metabolic fluxes after the temperature upshift were redirected similarly to the production strain; the effects, however, were observed to a lesser extent and with different time profiles. © 2002 Wiley Periodicals, Inc. Biotechnol Bioeng 80: 320–330, 2002.