Solubility enhancement of aggregation-prone heterologous proteins by fusion expression using stress-responsive Escherichia coli protein, RpoS.
BACKGROUND
The most efficient method for enhancing solubility of recombinant proteins appears to use the fusion expression partners. Although commercial fusion partners including maltose binding protein and glutathione-S-transferase have shown good performance in enhancing the solubility, they cannot be used for the proprietory production of commercially value-added proteins and likely cannot serve as universal helpers to solve all protein solubility and folding issues. Thus, novel fusion partners will continue to be developed through systematic investigations including proteome mining presented in this study.


RESULTS
We analyzed the Escherichia coli proteome response to the exogenous stress of guanidine hydrochloride using 2-dimensional gel electrophoresis and found that RpoS (RNA polymerase sigma factor) was significantly stress responsive. While under the stress condition the total number of soluble proteins decreased by about 7 %, but a 6-fold increase in the level of RpoS was observed, indicating that RpoS is a stress-induced protein. As an N-terminus fusion expression partner, RpoS increased significantly the solubility of many aggregation-prone heterologous proteins in E. coli cytoplasm, indicating that RpoS is a very effective solubility enhancer for the synthesis of many recombinant proteins. RpoS was also well suited for the production of a biologically active fusion mutant of Pseudomonas putida cutinase.


CONCLUSIONS
RpoS is highly effective as a strong solubility enhancer for aggregation-prone heterologous proteins when it is used as a fusion expression partner in an E. coli expression system. The results of these findings may, therefore, be useful in the production of other biologically active industrial enzymes, as successfully demonstrated by cutinase.
Park
Jin-Seung
JS
Department of Chemical and Biological Engineering, Korea University, Anam-Dong 5-1, Sungbuk-Ku, Seoul 136-713, South Korea. anorain@korea.ac.kr
Han
Kyung-Yeon
KY
Lee
Jong-Ho
JH
Song
Jong-Am
JA
Ahn
Keum-Young
KY
Seo
Hyuk-Seong
HS
Sim
Sang-Jun Jun
SJ
Kim
Seung-Wook
SW
Lee
Jeewon
J
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D019281:Dimerization; D004926:Escherichia coli / Q000502:physiology*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D018384:Oxidative Stress / Q000502:physiology; D011485:Protein Binding; D015202:Protein Engineering / Q000379:methods*; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D012808:Sigma Factor / Q000235:genetics / Q000378:metabolism*; D012995:Solubility
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D011993:Recombinant Fusion Proteins; D012808:Sigma Factor; C069567:sigma factor KatF protein, Bacteria