Mechanistic platform knowledge of concomitant sugar uptake in Escherichia coli BL21(DE3) strains

David J Wurm,Sophia Ulonska,Oliver Spadiut,Johanna Hausjell,Christoph Herwig

doi:10.1038/srep45072

David J Wurm, Sophia Ulonska + Show 3 more

Open Access

https://doi.org/10.1038/srep45072

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Abstract

When producing recombinant proteins, the use of Escherichia coli strain BL21(DE3) in combination with the T7-based pET-expression system is often the method of choice. In a recent study we introduced a mechanistic model describing the correlation of the specific glucose uptake rate (qs,glu) and the corresponding maximum specific lactose uptake rate (qs,lac,max) for a pET-based E. coli BL21(DE3) strain producing a single chain variable fragment (scFv). We showed the effect of qs,lac,max on productivity and product location underlining its importance for recombinant protein production. In the present study we investigated the mechanistic qs,glu/qs,lac,max correlation for four pET-based E. coli BL21(DE3) strains producing different recombinant products and thereby proved the mechanistic model to be platform knowledge for E. coli BL21(DE3). However, we found that the model parameters strongly depended on the recombinant product. Driven by this observation we tested different dynamic bioprocess strategies to allow a faster investigation of this mechanistic correlation. In fact, we succeeded and propose an experimental strategy comprising only one batch cultivation, one fed-batch cultivation as well as one dynamic experiment, to reliably determine the mechanistic model for qs,glu/qs,lac,max and get trustworthy model parameters for pET-based E. coli BL21(DE3) strains which are the basis for bioprocess development.

Highlights

The bacterium Escherichia coli is one of the most widely used host organisms for recombinant protein production[1,2,3,4]
To shed more light on the mechanistic correlation between the uptake of glucose and lactose, we recently performed a comprehensive study with a recombinant pET-based E. coli BL21(DE3) strain producing a single chain variable fragment against celiac disease[16]
We hypothesized that this mechanistic correlation might describe platform knowledge for E. coli BL21(DE3) strains carrying the pET expression system and proposed to conduct at

Summary

Introduction

The bacterium Escherichia coli is one of the most widely used host organisms for recombinant protein production[1,2,3,4]. Least four bioreactor cultivations (batch and fed-batch experiments) to determine the mechanistic model for any pET-based E. coli BL21(DE3) strain[16]. In the present study we put our hypothesis to test and investigated the mechanistic correlation of qs,glu and qs,lac,max for four pET-based E. coli BL21(DE3) strains producing different recombinant proteins.

Results

Conclusion