Solubility partner IF2 Domain i enables high yield synthesis of transducible transcription factors in Escherichia coli

William C. Yang, John P. Welsh, Jieun Lee, John P. Cooke, James R. Swartz

Research output: Contribution to journalArticle

7 Scopus citations

Abstract

Since the discovery that somatic cells could be reprogrammed back to a pluripotent state through the viral expression of a certain set of transcription factors, there has been great interest in reprogramming using a safer and more clinically relevant protein-based approach. However, the search for an efficient reprogramming approach utilizing the transcription factors in protein form requires a significant amount of protein material. Milligram quantities of transcription factors are challenging to obtain due to low yields and poor solubility. In this work, we describe enhanced production of the pluripotency transcription factors Oct4, Sox2, Klf4, Nanog, and Lin28 after fusing them to a solubility partner, IF2 Domain I (IF2D1). We expressed and purified milligram quantities of the fusion proteins. Though the transcription factor passenger proteins became insoluble after removal of the IF2D1, the un-cleaved Oct4, Sox2, Klf4, and Nanog fusion proteins exhibited specific binding to their consensus DNA sequences. However, when we administered the un-cleaved IF2D1-Oct4-R9 and IF2D1-Sox2-R9 to fibroblasts and measured their ability to influence transcriptional activity, we found that they were not fully bioactive; IF2D1-Oct4-R9 and IF2D1-Sox2-R9 influenced only a subset of their downstream gene targets. Thus, while the IF2D1 solubility partner enabled soluble production of the fusion protein at high levels, it did not yield fully bioactive transcription factors.

Original languageEnglish (US)
Pages (from-to)145-151
Number of pages7
JournalProtein Expression and Purification
Volume80
Issue number1
DOIs
StatePublished - Nov 2011

Keywords

  • IF2D1
  • iPSC
  • Pluripotency
  • Solubility partner
  • Transcription factor

ASJC Scopus subject areas

  • Biotechnology

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