Enhanced soluble protein expression using two new fusion tags.
Production of soluble recombinant proteins is vital for structure-function analysis and therapeutic applications. Unfortunately, when expressed in a heterologous host, such as Escherichia coli, most proteins are expressed as insoluble aggregates. Two new fusion partners have been identified to address these solubility problems. One of the tags was derived from a bacteriophage T7 protein kinase and the other one from a small E. coli chaperone, Skp. We have expressed a panel of insoluble human proteins including Hif1alpha, IL13, and folliculin as fusion proteins using these tags. Most of these fusion proteins were able to be expressed in a soluble form and could be purified by virtue of a Strep-tag II installed at the amino-terminal end of the fusion partners. In addition, we show that some of these proteins remained soluble after removal of the fusion tags by a site-specific protease. The results with these tags compare favorably to results with the most commonly used solubility tags described in the literature. Therefore, these two new fusion tags have the potential to express soluble proteins when fused with many recalcitrant proteins.
Chatterjee
Deb K
DK
Protein Expression Laboratory, SAIC-Frederick, Inc., National Cancer Institute at Frederick, 1050 Boyles Street, Building 327 Frederick, MD 21702, USA. chatterjee@ncifcrf.gov
Esposito
Dominic
D
D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods*; D017931:DNA Primers; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins / Q000235:genetics / Q000302:isolation & purification; D015723:Gene Library; D006801:Humans; D008969:Molecular Sequence Data; D010957:Plasmids; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification* / Q000378:metabolism; D015183:Restriction Mapping; D014764:Viral Proteins / Q000235:genetics / Q000302:isolation & purification
D017931:DNA Primers; D029968:Escherichia coli Proteins; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins