Detergent-solubilized Patched purified from Sf9 cells fails to interact strongly with cognate Hedgehog or Ihog homologs

Thomas E. Cleveland, Jacqueline M. McCabe, Daniel J. Leahy

Research output: Contribution to journalArticlepeer-review

Abstract

Patched (Ptc) is a twelve-pass transmembrane protein that functions as a receptor for the Hedgehog (Hh) family of morphogens. In addition to Ptc, several accessory proteins including the CDO/Ihog family of co-receptors are necessary for proper Hh signaling. Structures of Hh proteins bound to members of the CDO/Ihog family are known, but the nature of the full Hh receptor complex is not well understood. We have expressed the Drosophila Patched and Mouse Patched-1 proteins in Sf9 cells and find that Sonic Hedgehog will bind to Mouse Patched-1 in isolated Sf9 cell membranes but that purified, detergent-solubilized Ptc proteins do not interact strongly with cognate Hh and CDO/Ihog homologs. These results may reflect a nonnative conformation of detergent-solubilized Ptc or that an additional factor or factors lost during purification are required for high-affinity Ptc binding to Hh.

Original languageEnglish (US)
Pages (from-to)92-102
Number of pages11
JournalProtein Expression and Purification
Volume104
DOIs
StatePublished - Dec 2014

Keywords

  • Hedgehog signaling
  • Patched
  • Protein purification

ASJC Scopus subject areas

  • Biotechnology

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