Plasmid‐encoded protein: The principal factor in the “metabolic burden” associated with recombinant bacteria

Biotechnology and Bioengineering - Tập 35 Số 7 - Trang 668-681 - 1990
William E. Bentley1, N. Mirjalili1, Dana C. Andersen1, Robert H. Davis1, Dhinakar S. Kompala1
1Department of Chemical Engineering, UniversityofColorado, Boulder, Colorado 80309-0424

Tóm tắt

AbstractExperimental elucidation of the metabolic load placed on bacteria by the expression of foreign protein is presented. The host/vector system is Escherichia coli RR1/pBR329 (ampr, camr, and letr). Plasmid content results, which indicate that the plasmid copy number monotonically increases with decreasing growth rate, are consistent with the literature on ColE1‐like plasmids. More significantly, we have experimentally quantified the reduction in growth rate brought about by the expression of chloramphenicol‐acetyl‐transferase (CAT) and β‐lactamase. Results indicate a nearly linear decrease in growth rate with increasing foreign protein content. Also, the change in growth rate due to foreign protein expression depends on the growth rate of the cells. The observed linear relationship is media independent and, to our knowledge, previously undocumented. Furthermore, the induction of CAT, mediated by the presence of chloramphenicol, is shown to occur only at low growth rates, which further increases the metabolic load.Results are vdelineated with the aid of a structured kinetic model representing the metabolism of recombinant E. coli. In this article, several previous hypotheses and model predictions are justified and validated. This work provides an important step in the development of comprehensive, methabolically‐structured, kinetic models capable of prediciting optimal conditions for maximizing product yield.

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Biotechnology and Bioengineering

Tập 35 Số 7

668-681

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