Premium
Combined phospho‐ and glycoproteome enrichment in nephrocalcinosis tissues of phytate‐fed rats
Author(s) -
Tran TrangHuyen,
Park JongMoon,
Kim OkHee,
Kim Bora,
Choi Doyoung,
Lee JeongHwa,
Kim KwangPyo,
Oh ByungChul,
Lee Hookeun
Publication year - 2013
Publication title -
rapid communications in mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.528
H-Index - 136
eISSN - 1097-0231
pISSN - 0951-4198
DOI - 10.1002/rcm.6742
Subject(s) - chemistry , glycoproteomics , phosphorylation , chromatography , tandem mass tag , tandem mass spectrometry , proteomics , biochemistry , glycosylation , sample preparation , mass spectrometry , quantitative proteomics , gene
RATIONALE Protein post‐translational modifications (PTMs) are directly involved in protein function and cellular activities. Among them, glycosylation and phosphorylation are particularly important modifications on proteins located at extracellular and intracellular domains, respectively. However, the combined detection using phospho‐ and glycoproteomics is limited mainly due to protocol differences. METHODS In this study, we developed a novel method for both phospho‐ and glycoproteome detection from a single sample batch, in which a titanium dioxide cartridge was used to capture the phosphoproteome, and the flow‐through solution was processed for capturing N‐linked glycopeptides using hydrazide resin. RESULTS By using 1 mg of protein from kidney tissue lysates from normal and diseased rats, we concurrently identified 437 glycosites/358 phosphosites and 468 glycosites/369 phosphosites in normal and disease kidneys, respectively, by liquid chromatography/tandem mass spectrometric analysis. CONCLUSIONS Compared with individual PTM analyses, the combined PTM analysis clearly provides more broad implications for PTMs related to the pathological status and discovery of biomarker candidates. Furthermore, the combined protocol thoroughly showed its advantages in enrichment efficiency and biological interpretation compared with current methods. Copyright © 2013 John Wiley & Sons, Ltd.