Characterization and identification of the protein partners of Fn3 domain in FnTm2

Peter James Baker, Yan Mei Chan, Moritz Hertel, Jin Kim Montclare

Research output: Contribution to journalArticle

Abstract

Recently, a novel transmembrane protein was found to be up-regulated in the auditory learning pathway of birds and mammals. The protein, FnTm2, was predicted to have an extracellular fibronectin III (Fn3) domain and a single transmembrane domain. By contrast to other studied Fn3 domains the extracellular domain of FnTm2 bears several cysteine residues, which are predicted to form disulfide bonds. The Fn3 domain of the FnTm2 protein was expressed in DH5-α Escherichia coli (E. coli) cells, purified and characterized by circular dichroism (CD). In order to identify binding partners to Fn3, the isolated protein was incubated with bird brain lysate for a pull down treatment. Of the proteins recognized, myelin basic protein (MBP) was identified as a bona fide partner; it was further characterized for binding to Fn3 in vitro via fluorescence spectroscopy and confirmed via isothermal calorimetry (ITC).

Original languageEnglish (US)
Pages (from-to)42-48
Number of pages7
JournalProtein Expression and Purification
Volume81
Issue number1
DOIs
StatePublished - Jan 1 2012

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Keywords

  • Cell signaling
  • Fn3 domain
  • Protein expression
  • Protein purification
  • Protein-protein interactions
  • Refolding

ASJC Scopus subject areas

  • Biotechnology

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