A Multidimensional Chromatography Technology for In-depth Phosphoproteome Analysis

• DOI: 10.1074/mcp.m700468-mcp200
• Publication Date: 2008-04-12T00:27:54Z
• Authors: Claudio P. Albuqu...; Marcus B. Smolka; Samuel H. Payne; Vineet Bafna; Jimmy Eng; Huilin Zhou
• Keywords: Phosphopeptides; Proteomics; 0301 basic medicine; Chromatography; Proteome; Saccharomyces cerevisiae; Phosphoproteins; Models, Biological; 03 medical and health sciences; Cluster Analysis; Metabolic Networks and Pathways; DNA Damage; Protein Binding

Protein phosphorylation is a post-translational modification widely used to regulate cellular responses. Recent studies showed that global analysis could be study signaling pathways and identify targets of protein kinases in cells. A key objective obtain an in-depth mapping low abundance cells; this necessitates the use suitable separation techniques because complexity phosphoproteome. Here we developed multidimensional chromatography technology, combining IMAC, hydrophilic interaction chromatography, reverse phase LC, for phosphopeptide purification fractionation. Its application yeast Saccharomyces cerevisiae after DNA damage led identification 8764 unique phosphopeptides from 2278 phosphoproteins using tandem MS. Analysis two proteins, Rad9 Mrc1, revealed approximately 50% their was identified via analysis. Thus, technology suited phosphoproteome studies.