Screening for soluble expression constructs using cell-free protein synthesis

International Journal of Biological Macromolecules
T LamlaM M T Bauer

Abstract

The SH2 domain of STAT6 was chosen to test the in vitro protein synthesis as a screening tool. Goal of the screening was to obtain constructs which produce soluble protein in E. coli. The expression of 70 different constructs using an E. coli based cell-free system revealed two constructs, which give partly soluble protein. The introduction of two mutations, which had been suggested by a structural based alignment of 20 different SH2 domains lead to increased solubility. The expression of both constructs in E. coli followed by an affinity and size exclusion chromatography resulted in milligram quantities of highly purified protein.

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Citations

Mar 31, 2011·Future Medicinal Chemistry·Kate MaclaganRenos Savva
Sep 10, 2014·Protein Expression and Purification·Carolin RichterJürgen Hubbuch
May 29, 2013·The FEBS Journal·Raymond D Socha, Nobuhiko Tokuriki
May 28, 2010·Protein Science : a Publication of the Protein Society·Barbara MaertensFrank Schäfer
Jun 19, 2010·The Journal of Steroid Biochemistry and Molecular Biology·Edmund Maser, Guangming Xiong

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