Data-directed top-down Fourier-transform mass spectrometry of a large integral membrane protein complex

Photosystem II from Galdieria sulphuraria

Balakumar Thangaraj, Christopher M. Ryan, Puneet Souda, Kirsten Krause, Kym F. Faull, Andreas P M Weber, Petra Fromme, Julian P. Whitelegge

Research output: Contribution to journalArticle

29 Citations (Scopus)

Abstract

High-resolution top-down MS was used to characterize eleven integral and five peripheral subunits of the 750 kDa photosystem II complex from the eukaryotic red alga, Galdieria sulphuraria. The primary separation used LC MS with concomitant fraction collection (LC-MS+), yielding around 40 intact mass tags at 100ppm mass accuracy on a low-resolution ESI mass spectrometer, whose retention and mass were used to guide subsequent high-resolution topdown nano-electrospray FT ion-cyclotron resonance MS experiments (FT-MS). Both collisionally activated and electron capture dissociation were used to confirm the presence of eleven small subunits to mass accuracy within 5 ppm; PsbE, PsbF, PsbH, PsbI, PsbJ, PsbK, PsbL, PsbM, PsbT, PsbX and PsbZ. All subunits showed covalent modifications that fall into three classes including retention of initiating formyl-methionine, removal of methionine at the N-terminus with or without acetylation, and removal of a longer N-terminal peptide. Peripheral subunits identified by top-down analysis included oxygen-evolving complex subunits PsbO, PsbU, PsbV, as well as Psb28 (PsbW) and Psb27 ("PsbZ-like"). Top-down high-resolution MS provides the necessary precision, typically less than 5 ppm, for identification and characterization of polypeptide composition of these important membrane protein complexes.

Original languageEnglish
Pages (from-to)3644-3656
Number of pages13
JournalProteomics
Volume10
Issue number20
DOIs
Publication statusPublished - Oct 2010

Fingerprint

Photosystem II Protein Complex
Fourier Analysis
Methionine
Mass spectrometry
Mass Spectrometry
Fourier transforms
Membrane Proteins
Cyclotrons
Rhodophyta
Acetylation
Cyclotron resonance
Peptides
Mass spectrometers
Algae
Electrons
Ions
Oxygen
Chemical analysis
Experiments

Keywords

  • FT-MS
  • Membrane proteins
  • Photosystem II
  • Red algae
  • Technology
  • Top-down proteomics

ASJC Scopus subject areas

  • Molecular Biology
  • Biochemistry

Cite this

Thangaraj, B., Ryan, C. M., Souda, P., Krause, K., Faull, K. F., Weber, A. P. M., ... Whitelegge, J. P. (2010). Data-directed top-down Fourier-transform mass spectrometry of a large integral membrane protein complex: Photosystem II from Galdieria sulphuraria. Proteomics, 10(20), 3644-3656. https://doi.org/10.1002/pmic.201000190

Data-directed top-down Fourier-transform mass spectrometry of a large integral membrane protein complex : Photosystem II from Galdieria sulphuraria. / Thangaraj, Balakumar; Ryan, Christopher M.; Souda, Puneet; Krause, Kirsten; Faull, Kym F.; Weber, Andreas P M; Fromme, Petra; Whitelegge, Julian P.

In: Proteomics, Vol. 10, No. 20, 10.2010, p. 3644-3656.

Research output: Contribution to journalArticle

Thangaraj, Balakumar ; Ryan, Christopher M. ; Souda, Puneet ; Krause, Kirsten ; Faull, Kym F. ; Weber, Andreas P M ; Fromme, Petra ; Whitelegge, Julian P. / Data-directed top-down Fourier-transform mass spectrometry of a large integral membrane protein complex : Photosystem II from Galdieria sulphuraria. In: Proteomics. 2010 ; Vol. 10, No. 20. pp. 3644-3656.
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