Symposium Sessions (Day1, Day2, Day3)
Basic Sessions (Day1, Day2, Day3)
Young Researchers' Sessions (Day1, Day2, Day3)
Poster Presentations
- Day 2, May 16（Tue.） Room P (Foyer, Room 1004-1007)
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2P-08 PDF
Tyrosine Phosphorylated Peptide Enrichment by Ligand Exchange Chromatography
Reversible protein phosphorylation of serine, threonine, and tyrosine residues is an essential post-translational modification of intracellular signal transduction. Among these, tyrosine phosphorylation is known to regulate the activity of key molecules involved in disease pathogenesis and drug targeting, such as receptor-type tyrosine kinases, but its intracellular abundance is only a few tenths that of serine and threonine phosphorylation, making it difficult to identify and quantify it on a large scale. In this study, we developed a method for selectively enriching tyrosine phosphopeptides by ligand exchange chromatography. Phosphopeptides enriched by titania chromatography from pervanadate-treated HeLa cell digests were used as samples and loaded onto a pipette-tip micro column packed with Phos-tag agarose resin with copper ion (II). The elution conditions were optimized by analyzing the percentage of tyrosine phosphopeptides in the elution fractions by LC/MS/MS. As a result, the number of tyrosine phosphopeptides identified was more than three times greater than without the Cu2+-Phos-tag column. The proportion of acidic amino acids and multiply phosphorylated peptides increased in the later elution fractions, and the phosphosites on the peptides tended to shift from the N-termini to the internal region.