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Sulfo‐NHS‐SS‐biotin derivatization: A versatile tool for MALDI mass analysis of PTMs in lysine‐rich proteins
Author(s) -
Markoutsa Stavroula,
Bahr Ute,
Papasotiriou Dimitrios G.,
Häfner AnnKathrin,
Karas Michael,
Sorg Bernd L.
Publication year - 2014
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.201300309
Subject(s) - lysine , chemistry , derivatization , acetylation , biotin , mass spectrometry , biochemistry , chromatography , proteolysis , enzyme , amino acid , gene
The discovery of PTMs in proteins by MS requires nearly complete sequence coverage of the detected proteolytic peptides. Unfortunately, mass spectrometric analysis of the desired sequence fragments is often impeded due to low ionization efficiency and/or signal suppression in complex samples. When several lysine residues are in close proximity tryptic peptides may be too short for mass analysis. Moreover, modified peptides often appear in low stoichiometry and need to be enriched before analysis. We present here how the use of sulfo‐NHS‐SS‐biotin derivatization of lysine side chain can help to detect PTMs in lysine‐rich proteins. This label leads to a mass shift which can be adjusted by reduction of the SS bridge and alkylation with different reagents. Low intensity peptides can be enriched by use of streptavidin beads. Using this method, the functionally relevant protein kinase A phosphorylation site in 5‐lipoxygenase was detected for the first time by MS. Additionally, methylation and acetylation could be unambiguously determined in histones.