High cell density fermentation of recombinant Escherichia coli expressing human interferon alpha 1

SummaryA defined medium was developed which, by means of a specific fed-batch mode, allows growth of the recombinant Escherichia coli strain TG1 (pBB210) up to a cell density of 60 g dry weight/l. Apart from glucose and aqueous ammonia fed as carbon and nitrogen sources, it was unnecessary to supply other nutrients or O2-enriched air. Aqueous ammonia also served for pH control. The pO2 level was kept at 20% saturation via closed-loop controls operating the two output variables of stirrer speed and glucose feeding rate. This fed-batch method prevented significant accumulation of acetate and other metabolic by-products. The recombinant E. coli expressed interferon alpha 1 more efficiently at a lower specific growth rate (μPr ≈ 0.15 h−1) than at the maximum specific growth rate (μmax = 0.45 h−1). Therefore, fermentation in the batch phase at μmax was only allowed to continue up to a medium cell density. In the succeeding fed-batch phase, the specific growth rate was reduced to μPr by increasing the stirrer speed according to an empirically developed time scale.

[1]  J. Shiloach,et al.  High‐yield growth of E. coli at different temperatures in a bench scale fermentor , 1975 .

[2]  S. Bauer,et al.  Pilot scale exponential growth of Escherichia coli W to high cell concentration with temperature variation , 1976, Biotechnology and bioengineering.

[3]  R. Breitling,et al.  Secretory expression inEscherichia coli andBacillus subtilis of human interferon α genes directed by staphylokinase signals , 1989, Molecular and General Genetics MGG.

[4]  F. J. Bailey,et al.  High-cell-density fermentation studies of a recombinantE. coli that expresses atrial natriuretic factor , 1987, Journal of Industrial Microbiology.

[5]  P. Denéfle,et al.  High-cell density fermentation studies of recombinant Escherichia coli strains expressing human interleukin-1 beta. , 1988, Annales de l'Institut Pasteur. Microbiology.

[6]  S. Shimizu,et al.  Effect of amino acid supplement on cell yield and gene product in Escherichia coli harboring plasmid , 1986, Biotechnology and bioengineering.

[7]  P. Denéfle,et al.  High-cell density fermentation studies of recombinant Escherichia coli strains expressing human interleukin-1β , 1988 .

[8]  A. Fiechter.  Physical and chemical parameters of microbial growth , 1984 .

[9]  J. Rhee,et al.  Physiological constraints in increasing biomass concentration ofEscherichiacoli B in fed-batch culture , 1987, Biotechnology Letters.

[10]  Armin Fiechter,et al.  Mass culture of Escherichia coli: Medium development for low and high density cultivation of Escherichia coli B/r in minimal and complex media , 1985 .

[11]  S. J. Pirt,et al.  Principles of microbe and cell cultivation , 1975 .

[12]  J. Cutayar,et al.  High cell density culture of E. coli in a fed-batch system with dissolved oxygen as substrate feed indicator , 1989, Biotechnology Letters.

[13]  Shoichi Shimizu,et al.  HIGH DENSITY CULTIVATION OF BIOMASS IN FED-BATCH SYSTEM WITH DO-STAT , 1979 .

[14]  C. Cooney,et al.  Growth of Microorganisms , 2001 .

[15]  E. Ziv,et al.  Dense growth of aerobic bacteria in a bench‐scale fermentor , 1976, Biotechnology and bioengineering.

[16]  T. Ritch,et al.  Production of human alpha consensus interferon in recombinant Escherichia coli , 1986 .

[17]  J. Shiloach,et al.  Use of a novel air separation system in a fed-batch fermentative culture of Escherichia coli , 1989, Applied and environmental microbiology.

[18]  U. Rinas,et al.  Glucose as a substrate in recombinant strain fermentation technology , 1989, Applied Microbiology and Biotechnology.

[19]  B G Thompson,et al.  Control of ammonium concentration in Escherichia coli fermentations , 1985, Biotechnology and bioengineering.

[20]  G. Winter,et al.  Improved oligonucleotide site-directed mutagenesis using M13 vectors. , 1985, Nucleic acids research.

[21]  I. Gleiser,et al.  Growth of E. Coli W to high cell concentration by oxygen level linked control of carbon source concentration , 1981 .

[22]  J. Shiloach,et al.  Maximal exponential growth rate and yield of E. coli obtainable in a bench‐scale fermentor , 1974, Biotechnology and bioengineering.