Cloning, expression, and preliminary structural characterization of RTN-1C

Barbara Fazi, Sonia Melino, Federica Di Sano, Daniel O. Cicero, Mauro Piacentini, Maurizio Paci

Research output: Contribution to journalArticlepeer-review

Abstract

Reticulons (RTNs) are endoplasmic reticulum-associated proteins widely distributed in plants, yeast, and animals. They are characterized by unique N-terminal parts and a common 200 amino acid C-terminal domain containing two long hydrophobic sequences. Despite their implication in many cellular processes, their molecular structure and function are still largely unknown. In this study, the reticulon family member RTN-1C has been expressed and purified in Escherichia coli and its molecular structure has been analysed by fluorescence and CD spectroscopy in different detergents in order to obtain a good solubility and a relative stability. The isotopically enriched protein has been also produced to perform structural studies by NMR spectroscopy. The preliminary results obtained showed that RTN-1C protein possesses helical transmembrane segments when a membrane-like environment is produced by detergents. Moreover, fluorescence experiments indicated the exposure of tryptophan side chains as predicted by structure prediction programs. We also produced the isotopically labelled protein and the procedure adopted allowed us to plan future NMR studies to investigate the biochemical behaviour of reticulon-1C and of its peptides spanning out from the membrane.

Original languageEnglish
Pages (from-to)881-886
Number of pages6
JournalBiochemical and Biophysical Research Communications
Volume342
Issue number3
DOIs
Publication statusPublished - Apr 14 2006

Keywords

  • Endoplasmic reticulum
  • Nogo
  • Protein structural characterization
  • Reticulon

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Fingerprint Dive into the research topics of 'Cloning, expression, and preliminary structural characterization of RTN-1C'. Together they form a unique fingerprint.

Cite this