Proteomic analysis of the retinal rod outer segment disks

Isabella Panfoli, Luca Musante, Angela Bachi, Silvia Ravera, Daniela Calzia, Angela Cattaneo, Maurizio Bruschi, Paolo Bianchini, Alberto Diaspro, Alessandro Morelli, Isidoro M. Pepe, Carlo Tacchetti, Giovanni Candiano

Research output: Contribution to journalArticlepeer-review

Abstract

The initial events of vision at low light take place in vertebrate retinal rods. The rod outer segment consists of a stack of flattened disks surrounded by the plasma membrane. A list of the proteins that reside in disks has not been achieved yet. We present the first comprehensive proteomic analysis of purified rod disks, obtained by combining the results of two-dimensional gel electrophoresis separation of disk proteins to MALDI-TOF or nLC-ESI-MS/MS mass spectrometry techniques. Intact disks were isolated from bovine retinal rod outer segments by a method that minimizes contamination from inner segment. Out of a total of 187 excised spots, 148 proteins were unambiguously identified. An additional set of 61 proteins (partially overlapping with the previous ones) was generated by one-dimensional (1D) gel nLC-ESI-MS/MS method. Proteins involved in vision as well as in aerobic metabolism were found, among which are the five complexes of oxidative phosphorylation. Results from biochemical, Western blot, and confocal laser scanning microscopy immunochemistry experiments suggest that F1F0-ATP synthase is located and catalytically active in ROS disk membranes. This study represents a step toward a global physiological characterization of the disk proteome and provides information necessary for future studies on energy supply for phototransduction.

Original languageEnglish
Pages (from-to)2654-2669
Number of pages16
JournalJournal of Proteome Research
Volume7
Issue number7
DOIs
Publication statusPublished - Jul 2008

Keywords

  • Confocal laser scanning microscopy
  • Disks
  • FF-ATP synthase
  • MALDI-TOF
  • nLC-ESI-MS/MS
  • Redox chain complexes
  • Rod outer segment
  • Two-dimensional polyacrylamide gel electrophoresis

ASJC Scopus subject areas

  • Biochemistry
  • Chemistry(all)
  • Genetics
  • Biotechnology

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