Publication Information
Carroll et al., 2008
No external accession available
Abstract
Mol Cell Proteomics. 2008 Feb;7(2):347-69. doi: 10.1074/mcp.M700052-MCP200. Epub
2007 Oct 13.
Analysis of the Arabidopsis cytosolic ribosome proteome provides detailed
insights into its components and their post-translational modification.
Carroll AJ(1), Heazlewood JL, Ito J, Millar AH.
Author information:
(1)Australian Research Council (ARC) Centre of Excellence in Plant Energy
Biology and School of Biomedical, Biomolecular and Chemical Sciences, The
University of Western Australia, 35 Stirling Highway, M316, Crawley 6009,
Western Australia, Australia.
Finding gene-specific peptides by mass spectrometry analysis to pinpoint gene
loci responsible for particular protein products is a major challenge in
proteomics especially in highly conserved gene families in higher eukaryotes. We
used a combination of in silico approaches coupled to mass spectrometry analysis
to advance the proteomics insight into Arabidopsis cytosolic ribosomal
composition and its post-translational modifications. In silico digestion of all
409 ribosomal protein sequences in Arabidopsis defined the proportion of
theoretical gene-specific peptides for each gene family and highlighted the need
for low m/z cutoffs of MS ion selection for MS/MS to characterize low molecular
weight, highly basic ribosomal proteins. We undertook an extensive MS/MS survey
of the cytosolic ribosome using trypsin and, when required, chymotrypsin and
pepsin. We then used custom software to extract and filter peptide match
information from Mascot result files and implement high confidence criteria for
calling gene-specific identifications based on the highest quality unambiguous
spectra matching exclusively to certain in silico predicted gene- or gene
family-specific peptides. This provided an in-depth analysis of the protein
composition based on 1446 high quality MS/MS spectra matching to 795 peptide
sequences from ribosomal proteins. These identified peptides from five gene
families of ribosomal proteins not identified previously, providing experimental
data on 79 of the 80 different types of ribosomal subunits. We provide strong
evidence for gene-specific identification of 87 different ribosomal proteins
from these 79 families. We also provide new information on 30 specific sites of
co- and post-translational modification of ribosomal proteins in Arabidopsis by
initiator methionine removal, N-terminal acetylation, N-terminal methylation,
lysine N-methylation, and phosphorylation. These site-specific modification data
provide a wealth of resources for further assessment of the role of ribosome
modification in influencing translation in Arabidopsis.
DOI: 10.1074/mcp.M700052-MCP200
PMID: 17934214 [Indexed for MEDLINE]
