imprint:
Springer Science and Business Media LLC, 2013
Published in:Microbial Cell Factories
Language:
English
DOI:
10.1186/1475-2859-12-58
ISSN:
1475-2859
Origination:
Footnote:
Description:
<jats:title>Abstract</jats:title>
<jats:sec>
<jats:title>Background</jats:title>
<jats:p>In the biopharmaceutical industry, <jats:italic>Escherichia coli</jats:italic> (<jats:italic>E. coli</jats:italic>) strains are among the most frequently used bacterial hosts for producing recombinant proteins because they allow a simple process set-up and they are Food and Drug Administration (FDA)-approved for human applications. Widespread use of <jats:italic>E. coli</jats:italic> in biotechnology has led to the development of many different strains, and selecting an ideal host to produce a specific protein of interest is an important step in developing a production process. <jats:italic>E. coli</jats:italic> B and K–12 strains are frequently employed in large-scale production processes, and therefore are of particular interest. We previously evaluated the individual cultivation characteristics of <jats:italic>E. coli</jats:italic> BL21 and the K–12 hosts RV308 and HMS174. To our knowledge, there has not yet been a detailed comparison of the individual performances of these production strains in terms of recombinant protein production and system stability. The present study directly compared the T7-based expression hosts <jats:italic>E. coli</jats:italic> BL21(DE3), RV308(DE3), and HMS174(DE3), focusing on evaluating the specific attributes of these strains in relation to high-level protein production of the model protein recombinant human superoxide dismutase (SOD). The experimental setup was an exponential carbon-limited fed-batch cultivation with minimal media and single-pulse induction.</jats:p>
</jats:sec>
<jats:sec>
<jats:title>Results</jats:title>
<jats:p>The host strain BL21(DE3) produced the highest amounts of specific protein, followed by HMS174(DE3) and RV308(DE3). The expression system HMS174(DE3) exhibited system stability by retaining the expression vector over the entire process time; however, it entirely stopped growing shortly after induction. In contrast, BL21(DE3) and RV308(DE3) encountered plasmid loss but maintained growth. RV308(DE3) exhibited the lowest ppGpp concentration, which is correlated with the metabolic stress level and lowest degradation of soluble protein fraction compared to both other strains.</jats:p>
</jats:sec>
<jats:sec>
<jats:title>Conclusions</jats:title>
<jats:p>Overall, this study provides novel data regarding the individual strain properties and production capabilities, which will enable targeted strain selection for producing a specific protein of interest. This information can be used to accelerate future process design and implementation.</jats:p>
</jats:sec>