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http://dx.doi.org/10.17480/psk.2015.59.3.113

A Comparison between C4 and Cation-exchange Columns as a Pre-separation Method for Mass Spectrometric Analysis to Characterize a Global Identification of Phosphopeptides and Phosphorylation Sites

Kim, Hye-Jeong (Department of Molecular Medicine, School of Medicine, Cell and Matrix Research Institute, Kyungpook National University)
Baek, Moon-Chang (Department of Molecular Medicine, School of Medicine, Cell and Matrix Research Institute, Kyungpook National University)

Publication Information

YAKHAK HOEJI / v.59, no.3, 2015 , pp. 113-119 More about this Journal

Abstract

Protein phosphorylation is one of most important post-translational modifications (PTMs) and plays an important role in regulation of protein function. Here we develop a method for a global identification of phosphopeptides and phosphorylation sites using nano-LC MS/MS. We compared two separation methods, C4 and strong cation ion exchange (SCX). Before phosphopeptides enrichment with $TiO_2$ , total proteins from Rat 1 cells have been separated using C4 column or tryptic peptides of proteins from the cells have been separated using SCX column. Finally, we have detected 52 phosphorylation sites on 41 proteins from SCX method and 375 phosphorylation sites on 252 proteins from C4 method, and determined the function and localization of identified phosphoproteins using DAVID software. In particular, we showed new phosphorylation sites from membrane proteins related to various cell signaling mechanisms. This method may contribute to study global signal networks induced by various signals including ligands and drugs.

Keywords

phosphopeptide; mass spectrometry; strong cation ion exchange; C4 column; $TiO_2$ ;

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