Preparation of bioactive soluble human leukemia inhibitory factor from recombinant Escherichia coli using thioredoxin as fusion partner.
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Authors
Tomala, MagdaLavrentieva, Antonina
Moretti, Pierre
Rinas, Ursula
Kasper, Cornelia
Stahl, Frank
Schambach, Axel
Warlich, Eva
Martin, Ulrich
Cantz, Tobias
Scheper, Thomas
Issue Date
2010-09
Metadata
Show full item recordAbstract
Leukemia inhibitory factor (LIF) is a polyfunctional cytokine with numerous regulatory effects in vivo and in vitro. In stem cell cultures it is the essential media supplement for the maintenance of pluripotency of embryonic and induced pluripotent stem cells. With regard to large scale cultures of these cells, LIF is needed in high quality and quantity and represents the major cost determining factor (90%) of the culture media. In this report, we describe a novel production and purification process for human LIF (hLIF) from recombinant Escherichia coli cultures. hLIF was cloned into pET32b and expressed as soluble protein in fusion with thioredoxin. After purification based on membrane adsorber technology, the fusion protein was cleaved using TEV protease. Released, soluble hLIF was subsequently purified by cation exchange chromatography and successfully tested for its biological activity using suspension cultures of murine embryonic and induced pluripotent stem cells. Our novel protocol for the production of recombinant hLIF is very suitable and effective for the production of poorly soluble proteins through expression in fusion with the solubilizing partner thioredoxin.Citation
Preparation of bioactive soluble human leukemia inhibitory factor from recombinant Escherichia coli using thioredoxin as fusion partner. 2010, 73 (1):51-7 Protein Expr. Purif.Affiliation
Institute of Technical Chemistry, Leibniz University of Hannover, Callinstr. 5, 30167 Hannover, Germany.PubMed ID
20381622Type
ArticleLanguage
enISSN
1096-0279ae974a485f413a2113503eed53cd6c53
10.1016/j.pep.2010.04.002
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