Analysis of
Protein Phosphorylation by Combination of IMAC, Phosphatase with Biological
Mass Spectrometry
WANG
Jing-Lan, ZHANG Yang-Jun, CAI Yun, LI Lei, LIU Shang-Yi, YANG He-Yi, QIAN
Xiao-Hong*
( Beijing Institute of Radiation Medicine, Beijing 100850,
China )
Abstract Protein phosphorylation is the most
important reversible post-translational modification in cells. Analysis of
phosphorylated proteins and identification of their phosphorylation sites is
helpful for understanding their biological functions. MALDI-TOF-MS and
ESI-Q-TOF-MS play important roles in protein phosphorylation analysis. In this
work, immobilized metal affinity chromatography (IMAC) was used to selectively
enrich phosphopeptides from protein digest mixtures, and treatment of
phosphopeptides with alkaline phosphatase was used to confirm the
phosphorylation. Finally, the phosphorylation sites were determined by tandem
mass spectrometry analysis and database searching.
Key words biological mass spectrometry;
phosphorylation; immobilized metal affinity chromatography
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