Abstract
BackgroundRecombinant protein production can be stressful to the host organism. The extent of stress is determined by the specific properties of the recombinant transcript and protein, by the rates of transcription and translation, and by the environmental conditions encountered during the production process.ResultsThe impact of the transcription of the T7-promoter controlled genes encoding human basic fibroblast growth factor (hFGF-2) and green fluorescent protein (GFP) as well as the translation into the recombinant protein on the growth properties of the production host E. coli BL21(DE3) were investigated. This was done by using expression vectors where the promoter region or the ribosome binding site(s) or both were removed. It is shown that already transcription without protein translation imposes a metabolic burden on the host cell. Translation of the transcript into large amounts of a properly folded protein does not show any effect on cell growth in the best case, e.g. high-level production of GFP in Luria–Bertani medium. However, translation appears to contribute to the metabolic burden if it is connected to protein folding associated problems, e.g. inclusion body formation.ConclusionThe so-called metabolic burden of recombinant protein production is mainly attributed to transcription but can be enhanced through translation and those processes following translation (e.g. protein folding and degradation, heat-shock responses).
Highlights
Recombinant protein production can be stressful to the host organism
High level recombinant protein production in Escherichia coli frequently leads to an impact on host cell metabolism detectable through
The factors contributing to the metabolic burden are multifaceted depending on the host/vector combination and on the properties of the encoded gene with its transcription and translation products as well as on the environmental conditions encountered during the production process [3,4,5,6]
Summary
The extent of stress is determined by the specific properties of the recombinant transcript and protein, by the rates of transcription and translation, and by the environmental conditions encountered during the production process. The most common E. coli-based expression system is the bacteriophage T7 RNA polymerase and T7 promoter controlled gene expression combination. T7 RNA polymerase leads to high level expression of T7 promoter-controlled genes and correspondingly high recombinant protein titers. High level recombinant protein production in Escherichia coli frequently leads to an impact on host cell metabolism detectable through. The factors contributing to the metabolic burden are multifaceted depending on the host/vector combination and on the properties of the encoded gene with its transcription and translation products as well as on the environmental conditions encountered during the production process [3,4,5,6]. Production of the same protein under identical conditions in the same host/vector combination but with single codon exchanges can change the metabolic perturbations considerably [10,11,12,13]
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