1. Accurate phosphorylation site localization using phospho-brackets.
- Author
-
Xiao K, Shen Y, Li S, and Tian Z
- Subjects
- Chemistry Techniques, Synthetic, Chromatography, Liquid, HeLa Cells, Humans, Peptide Library, Tandem Mass Spectrometry, Phosphopeptides chemistry, Phosphorylation, Proteomics
- Abstract
Phosphorylation is one of the most important and widely studied protein post-translational modifications. Tandem mass spectrometry using higher-energy collisional dissociation has evolved into a state-of-the-art analytical platform for both phosphorylation identification and site localization. Tens of thousands of phosphopeptides can now be routinely identified from a single shotgun proteomics study; site localization, however, is much more complicated and many challenges still exist. Here, we report our development of P-bracket using direct experimental evidence of phospho-containing site-determining product ions for accurate site localization without the need for additional FLR control. A P-bracket is defined as a complementary product ion pair that forms a bracket to confine a phosphorylation event to a unique site. P-bracket has been successfully benchmarked with a set of six synthetic phosphopeptides with a single phosphorylation event, a set of 96 synthetic peptides and phosphopeptide reference libraries, and two HeLa phosphopeptide LC-MS/MS (HCD) datasets; Accurate phosphosite localization by P-bracket will greatly enhance identification confidence of phosphopeptides and contribute to structural and functional studies of phosphoproteins., (Copyright © 2017 Elsevier B.V. All rights reserved.)
- Published
- 2017
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