Identifying phosphopeptide by searching a site annotated protein database

doi:10.3724/SP.J.1123.2014.10019

Abstract

Abstract:

Phosphoproteome analysis is one of the important research fields in proteomics. In shotgun proteomics, phosphopeptides could be identified directly by setting phosphorylation as variable modifications in database search. However, search space increases significantly when variable modifications are set in post-translation modifications (PTMs) analysis, which will decrease the identification sensitivity. Because setting a variable modification on a specific type of amino acid residue means all of this amino acid residues in the database might be modified, which is not consistent with actual conditions. Phosphorylation and dephosphorylation are regulated by protein kinases and phosphatases, which can only occur on particular substrates. Therefore only residues within specific sequence are potential sites which may be modified. To address this issue, we extracted the characteristic sequence from the identified phosphorylation sites and created an annotated database containing phosphorylation site information, which allowed the searching engine to set variable modifications only on the serine, threonine and tyrosine residues that were identified to be phosphorylated previously. In this database only annotated serine, threonine and tyrosine can be modified. This strategy significantly reduced the search space. The performance of this new database searching strategy was evaluated by searching different types of data with Mascot, and higher sensitivity for phosphopeptide identification was achieved with high reliability.

Key words: phosphorylation, proteomics, search space, site annotation

CLC Number:

O658

CHENG Kai, WANG Fangjun, BIAN Yangyang, YE Mingliang, ZOU Hanfa. Identifying phosphopeptide by searching a site annotated protein database[J]. Chinese Journal of Chromatography, 2015, 33(1): 10-16.

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