Document Detail


Identifying and quantifying in vivo methylation sites by heavy methyl SILAC.
MedLine Citation:
PMID:  15782174     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Protein methylation is a stable post-translational modification (PTM) with important biological functions. It occurs predominantly on arginine and lysine residues with varying numbers of methyl groups, such as mono-, di- or trimethyl lysine. Existing methods for identifying methylation sites are laborious, require large amounts of sample and cannot be applied to complex mixtures. We have previously described stable isotope labeling by amino acids in cell culture (SILAC) for quantitative comparison of proteomes. In heavy methyl SILAC, cells metabolically convert [(13)CD(3)]methionine to the sole biological methyl donor, [(13)CD(3)]S-adenosyl methionine. Heavy methyl groups are fully incorporated into in vivo methylation sites, directly labeling the PTM. This provides markedly increased confidence in identification and relative quantitation of protein methylation by mass spectrometry. Using antibodies targeted to methylated residues and analysis by liquid chromatography-tandem mass spectrometry, we identified 59 methylation sites, including previously unknown sites, considerably extending the number of in vivo methylation sites described in the literature.
Authors:
Shao-En Ong; Gerhard Mittler; Matthias Mann
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Publication Detail:
Type:  Evaluation Studies; Journal Article; Research Support, Non-U.S. Gov't; Validation Studies     Date:  2004-10-21
Journal Detail:
Title:  Nature methods     Volume:  1     ISSN:  1548-7091     ISO Abbreviation:  Nat. Methods     Publication Date:  2004 Nov 
Date Detail:
Created Date:  2005-05-03     Completed Date:  2005-05-24     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  101215604     Medline TA:  Nat Methods     Country:  United States    
Other Details:
Languages:  eng     Pagination:  119-26     Citation Subset:  IM    
Affiliation:
Center for Experimental BioInformatics, University of Southern Denmark, Odense M 5230, Denmark.
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MeSH Terms
Descriptor/Qualifier:
Algorithms*
Binding Sites
Carbon Isotopes
Chromatography, Liquid / methods*
Gene Expression Profiling / methods
Hela Cells
Humans
Isotope Labeling / methods*
Mass Spectrometry / methods*
Methionine / analysis,  metabolism*
Methylation
Neoplasm Proteins / analysis,  metabolism*
Protein Binding
Proteome / analysis,  metabolism*
Chemical
Reg. No./Substance:
0/Carbon Isotopes; 0/Neoplasm Proteins; 0/Proteome; 63-68-3/Methionine

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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