Mass Spectrometric Characterization of Human Histone H3: A Bird's Eye View

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Abstract
The modification of H3 in asynchronous HeLa cells was profiled using Top Down Mass Spectrometry. A broad distribution of species differing by 14 Da and containing less than 3% unmodified protein was observed for all three variants. Species of up to +168 Da were observed for H3.1, and fragmentation of all species by Electron Capture Dissociation (ECD) revealed ∼5% methylation of K4 and ∼50% dimethylation of K9. K14 and K23 were major sites of acetylation. H3.3 was slightly hypermodified with the apex of the distribution shifted by ∼+14 Da compared to H3.1. H3.1 (50% and 15%) from colchicine-treated cells was monophosphorylated and diphosphorylated, respectively, with equivalent modification of S10 and S28. Keywords: H3 • histone code • histone • Fourier transform mass spectrometry (FTMS) • electron capture dissociation (ECD) • chromatin • post-translational modification (PTM) • acetylation • methylation • phosphorylation