Large-Scale Identification of the Arginine Methylome by Mass Spectrometry
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Large-Scale Identification of the Arginine Methylome by Mass Spectrometry. / Sylvestersen, Kathrine B; Nielsen, Michael L.
Current Protocols in Protein Science. Vol. 82 2015. (Current Protocols in Protein Science).Research output: Chapter in Book/Report/Conference proceeding › Book chapter › Research › peer-review
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TY - CHAP
T1 - Large-Scale Identification of the Arginine Methylome by Mass Spectrometry
AU - Sylvestersen, Kathrine B
AU - Nielsen, Michael L
N1 - Copyright © 2015 John Wiley & Sons, Inc.
PY - 2015/11/2
Y1 - 2015/11/2
N2 - The attachment of one or more methylation groups to the side chain of arginine residues is a regulatory mechanism for cellular proteins. Recent advances in mass spectrometry-based characterization allow comprehensive identification of arginine methylation sites by peptide-level enrichment strategies. Described in this unit is a 4-day protocol for enrichment of arginine-methylated peptides and subsequent identification of thousands of distinct sites by mass spectrometry. Specifically, the protocol explains step-by-step sample preparation, enrichment using commercially available antibodies, prefractionation using strong cation exchange, and identification using liquid chromatography coupled to tandem mass spectrometry. A strategy for relative quantification is described using stable isotope labeling by amino acids in cell culture (SILAC). Approaches for analysis of arginine methylation site occupancy are also discussed. Collectively, the unit describes the essential parameters required for a successful and comprehensive experiment detailing the arginine methylome. © 2015 by John Wiley & Sons, Inc.
AB - The attachment of one or more methylation groups to the side chain of arginine residues is a regulatory mechanism for cellular proteins. Recent advances in mass spectrometry-based characterization allow comprehensive identification of arginine methylation sites by peptide-level enrichment strategies. Described in this unit is a 4-day protocol for enrichment of arginine-methylated peptides and subsequent identification of thousands of distinct sites by mass spectrometry. Specifically, the protocol explains step-by-step sample preparation, enrichment using commercially available antibodies, prefractionation using strong cation exchange, and identification using liquid chromatography coupled to tandem mass spectrometry. A strategy for relative quantification is described using stable isotope labeling by amino acids in cell culture (SILAC). Approaches for analysis of arginine methylation site occupancy are also discussed. Collectively, the unit describes the essential parameters required for a successful and comprehensive experiment detailing the arginine methylome. © 2015 by John Wiley & Sons, Inc.
U2 - 10.1002/0471140864.ps2407s82
DO - 10.1002/0471140864.ps2407s82
M3 - Book chapter
C2 - 26521714
VL - 82
T3 - Current Protocols in Protein Science
BT - Current Protocols in Protein Science
ER -
ID: 176889772