Relative Quantification of Membrane Proteins in Wild-Type and Prion Protein (PrP)-Knockout Cerebellar Granule Neurons

Research output: Contribution to journalArticle

Abstract

Approximately 25% of eukaryotic proteins possessing homology to at least two trans membrane domains are predicted to be embedded in biological membranes. Nevertheless, this group of proteins is not usually well represented in proteome-wide experiments due to their refractory nature. Here we present a quantitative mass spectrometry-based comparison of membrane protein expression in cerebellar granule neurons grown in primary culture that were isolated from wild-type mice and mice lacking the cellular prion protein. This protein is a cell-surface glycoprotein that is mainly expressed in the central nervous system and is involved in several neurodegenerative disorders, though its physiological role is unclear. We used a low specificity enzyme a-chymotrypsin to digest membrane proteins preparations that had been separated by SDS-PAGE. The resulting peptides were labeled with tandem mass tags and analyzed by MS. The differentially expressed proteins identified using this approach were further analyzed by multiple reaction monitoring to confirm the expression level changes.

Details

Authors
  • Roberto Stella
  • Paolo Cifani
  • Caterina Peggion
  • Karin M Hansson
  • Cristian Lazzari
  • Maria Bendz
  • Fredrik Levander
  • Maria Catia Sorgato
  • Alessandro Bertoli
  • Peter James
Organisations
Research areas and keywords

Subject classification (UKÄ) – MANDATORY

  • Immunology in the medical area
  • Health Sciences

Keywords

  • tandem mass tags, multiple reaction monitoring, mass spectrometry, gene knockout, membrane proteins, Prion protein, PrP
Original languageEnglish
Pages (from-to)523-536
JournalJournal of Proteome Research
Volume11
Issue number2
Publication statusPublished - 2012
Publication categoryResearch
Peer-reviewedYes