Site-specific quantification of lysine acetylation in the N-terminal tail of histone H4 using a double-labelling, targeted UHPLC MS/MS approach.

ABSTRACT: We developed a targeted liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the site-specific quantification of lysine acetylation in the N-terminal region of histone H4 by combining chemical derivatization at the protein and peptide levels with digestion using chymotrypsin and trypsin. Unmodified ε-amino groups were first modified with propionic acid anhydride and the derivatized protein digested with trypsin and chymotrypsin. The newly formed peptide N-termini were subjected to a second derivatization step with d6- (heavy) or d0- (light) acetic acid anhydride. Samples were mixed at different ratios and peptides monitored by multiple reaction monitoring (MRM) LC-MS/MS. The method was validated in terms of linearity (R(2) ≥ 0.94), precision (RSD ≤ 10%), and accuracy (≤27%)

SUBMITTER: D'Urzo A 

PROVIDER: S-EPMC4837199 | biostudies-literature | 2016 May

REPOSITORIES: biostudies-literature