Dynamics of induced CAT expression in E. coli

The dynamics of chemically induced chloramphenicolaceyl‐transferase (CAT) expression are determined in batch cultures of Escherichia coli DH5αF'IQ [pKK262‐1]. This article is directed towards understanding the coupling of induced cloned‐protein synthesis and reduced cell growth which are balanced in the optimal system. Experimental results indicate that the best inducer (IPTG) concentration is near 1.0 mM when added during midexponential growth. Lower concentrations cause only weak induction, whereas higher concentrations cause sufficiently strong induction that cell growth is suppressed. Induction at the onset of the stationary phase results in high expression but is accompanied by stimulated protease activity. Also, cell mass yield is adversely affected by enhanced protein synthesis. A structured metabolic model is shown to predict the responses of instantaneous growth rate and productivity which result from protein overexpression. This model can be employed to predict alternative reactor strategies and operating conditions necessary for the design of efficient bioprocess.

[1]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[2]  Robert H. Davis,et al.  Plasmid maintenance and protein overproduction in selective recycle bioreactors , 1991, Biotechnology and bioengineering.

[3]  Prasad Dhurjati,et al.  A comparison of mathematical model predictions to experimental measurements for growth and recombinant protein production in induced cultures of Escherichia coli , 1990, Biotechnology and bioengineering.

[4]  W. Bentley,et al.  Plasmid‐encoded protein: The principal factor in the “metabolic burden” associated with recombinant bacteria , 1990, Biotechnology and bioengineering.

[5]  T. Park,et al.  Optimization of fermentation processes using recombinant Escherichia coli with the cloned trp operon , 1989, Biotechnology and bioengineering.

[6]  K. Turner,et al.  Effect of induction temperature on the production of malaria antigens in recombinant E. coli , 1989, Biotechnology and bioengineering.

[7]  D. Kompala,et al.  A structured kinetic modeling framework for the dynamics of hybridoma growth and monoclonal antibody production in continuous suspension cultures , 1989, Biotechnology and bioengineering.

[8]  S. J. Parulekar,et al.  Expression of β‐lactamase by recombinant Escherichia coli strains containing plasmids of different sizes—effects of pH, phosphate, and dissolved oxygen , 1989, Biotechnology and bioengineering.

[9]  P. Dhurjati,et al.  Effects of plasmid amplification and recombinant gene expression on the growth kinetics of recombinant E. coli , 1989, Biotechnology and bioengineering.

[10]  C. W. Robinson,et al.  Induction of T4 DNA ligase in a recombinant strain of Escherichia coli , 1989, Biotechnology and bioengineering.

[11]  B O Palsson,et al.  On the kinetics of plasmid replication. , 1989, Journal of theoretical biology.

[12]  W. Ramirez,et al.  Dynamics of foreign protein secretion from Saccharomyces cerevisiae , 1989, Biotechnology and bioengineering.

[13]  W. Bentley,et al.  A novel structured kinetic modeling approach for the analysis of plasmid instability in recombinant bacterial cultures , 1989, Biotechnology and bioengineering.

[14]  G. Georgiou,et al.  Release of periplasmic enzymes and other physiological effects of β‐lactamase overproduction in Escherichia coli , 1988, Biotechnology and bioengineering.

[15]  J. Bailey,et al.  Transient response simulations of recombinant microbial populations , 1988, Biotechnology and bioengineering.

[16]  M. Tsuchiya,et al.  Control of Folding of Proteins Secreted by a High Expression Secretion Vector, pIN-III-ompA: 16-Fold Increase in Production of Active Subtilisin E in Escherichia Coli , 1988, Bio/Technology.

[17]  J. Wienberg,et al.  Amplified expression constructs for human tissue-type plasminogen activator in Chinese hamster ovary cells: instability in the absence of selective pressure. , 1988, Gene.

[18]  A. Goldberg,et al.  An increased content of protease La, the lon gene product, increases protein degradation and blocks growth in Escherichia coli. , 1987, The Journal of biological chemistry.

[19]  D. Zabriskie,et al.  Factors influencing productivity of fermentations employing recombinant microorganisms , 1986 .

[20]  F. Marston.  The purification of eukaryotic polypeptides synthesized in Escherichia coli. , 1986, The Biochemical journal.

[21]  J. Bailey,et al.  Theoretical growth yield estimates for recombinant cells. , 1986, Biotechnology and bioengineering.

[22]  J E Bailey,et al.  A kinetic model for product formation in unstable recombinant populations , 1985, Biotechnology and bioengineering.

[23]  M M Ataai,et al.  Simulation of CFSTR through development of a mathematical model for anaerobic growth of Escherichia coli cell population , 1985, Biotechnology and bioengineering.

[24]  J. Brosius,et al.  Spacing of the -10 and -35 regions in the tac promoter. Effect on its in vivo activity. , 1985, The Journal of biological chemistry.

[25]  R. Siegel,et al.  Kinetic study of instability of recombinant plasmid pPLc23trpAl in E. coli using two‐stage continuous culture system , 1985, Biotechnology and bioengineering.

[26]  J E Bailey,et al.  Genetically structured models forlac promoter–operator function in the Escherichia coli chromosome and in multicopy plasmids: Lac operator function , 1984, Biotechnology and bioengineering.

[27]  M M Domach,et al.  A finite representation model for an asynchronous culture of E. coli , 1984, Biotechnology and bioengineering.

[28]  J. Brosius,et al.  Vectors bearing a hybrid trp-lac promoter useful for regulated expression of cloned genes in Escherichia coli. , 1983, Gene.

[29]  R. Rodriguez,et al.  Recombinant DNA Techniques: An Introduction , 1983 .

[30]  W. Vieth,et al.  Active transport of inducer in enzyme biosynthesis. , 1982, Biotechnology and bioengineering.

[31]  A Constantinides,et al.  Modeling the role of cyclic AMP in catabolite repression of inducible enzyme biosynthesis in mircobial cells , 1978, Biotechnology and bioengineering.

[32]  T. Imanaka,et al.  A kinetic model of catabolite repression in the dual control mechanism in microorganisms. , 1977, Biotechnology and bioengineering.

[33]  K. Toda,et al.  Dual control of invertase Biosynthesis in chemostat culture , 1976, Biotechnology and bioengineering.

[34]  D. E. Brown.  Membrane Systems: Analysis and Design : By W. R. Vieth. Carl Hanser Verlag. Munchen. xvii + 360 pp. ISBN 3 446 15206 7. Hardback price £35.90. Oxford University Press, New York. ISBN 0 19 520767 X. , 1990 .

[35]  F. Neidhardt,et al.  Physiology of the bacterial cell : a molecular approach , 1990 .

[36]  S. Maeda.  Expression of foreign genes in insects using baculovirus vectors. , 1989, Annual review of entomology.