TECHNICAL NOTE
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Identification of a suitable host strain
When expressing a protein in an organism other than
its native host, a variety of issues can arise. Genetically
engineered E. coli strains with enhanced capabilities that
overcome toxicity or reduced translational efficiency are
available. Three such strains were tested, and the resulting
lysates compared to find the best host for His-tagged
HRV3C. All three strains were grown overnight at 20 °C in
TB medium with 0.5 mM IPTG before harvest and lysis.
Analysis of the clarified lysates in SDS-PAGE didn't reveal
a significant difference in total protein levels (Figure
4A). In contrast, analysis of the clarified lysates with
Andromeda suggests differences in the yield of folded
His-tagged HRV3C amongst the three strains, with strain
3 performing best (Figure 4B).
First
Derivative
Temperature [°C]
30 35 40 45 50 55 60
Strain 1
Strain 2
Strain 3
Figure 4: Samples collected from three different E. coli strains grown at identical conditions were analyzed via SDS-PAGE and Andromeda. A) SDS-PAGE of the lysate
(L), clarified lysate (C), and purified protein (P). The expected molecular weight of the protein of interest is indicated by an arrow. B) Andromeda unfolding profiles of
clarified lysate samples collected from strain 1 (purple), strain 2 (blue), and strain 3 (yellow). Unfolding profiles represent the average of at least three technical repli-
cates. The shaded area indicates the standard deviation amongst all technical replicates.
A B
