Modeling of the controlled expression of a harmful protein by a three‐plasmid harboring system

Abstract

A genetically structured mathematical model was developed and used to evaluate the influence of molecular parameters involved in the expression of a harmful recombinant protein (SPA::EcoRI). The system consists of the controlled expression of the endonuclease EcoRI cloned in the plasmid pMTC48. The control is exerted by the lambda CI repressor expressed from the plasmid pRK248cIts. The deleterious effect of the activity of the enzyme EcoRI on the host DNA is prevented by the action of the EcoRI methylase that is expressed constitutively from a third plasmid, pEcoR4. The model includes molecular mechanisms involved in the regulation of the expression of these genes and is used to determine cultural conditions that maximize the production of the recombinant protein.

Process Development in Biotechnology - A Re-Evaluation

“…The gene expression was performed in the second stage at 40 C or with IPTG addition and a protein supplement. A genetic structure model was developed for this process [8], which allows the regulation of the gene expression to be simulated. The methylase gene was expressed by the constitutive promoter.…”

“…a) Concentration of repression protein CI and restriction endonuclease EcoRI in recombinant E. coli at 42 C as a function of the doubling time[8]. b) Activity of restriction endonuclease EcoRI and methylase in recombinant E. coli at 30 C and 42 C as a function of the doubling time[8]. (Reprinted with permission of Wiley-Liss, Inc., a subsidiary of John Wiley & Sons, Inc.).…”

Prozessentwicklung in der Biotechnologie – Ein Rückblick

“…Ein genetisch strukturiertes mathematisches Modell wurde entwickelt [8], das die Bestimmung der Regulation der Genexpression ermöglicht. Das Methylase-Gen wird durch den konstitutiven Promotor exprimiert.…”