Ultra-High Performance Liquid Chromatography−Mass Spectrometry for the Fast Profiling of Histone Post-Translational Modifications

Histones are subjected to extensive post-translational modifications (PTMs) that are known to play key roles in many biological processes. In this study, we report a fast, efficient, highly reproducible, and easily automated method involving ultra-high performance liquid chromatography (UHPLC) coupled to a high resolution/high mass accuracy LTQ-Orbitrap mass spectrometer to profile core histone modifications/variants from WI-38 primary human fibroblasts. The whole analysis was performed on intact unfractionated histones within 19 min, which is ∼3-fold faster than previously published procedures. High mass accuracy measurements combined with top-down tandem mass spectrometry (MS) experiments enable accurate histone identification. Experimental and biological variations were thoroughly assessed and were 8% and 16% on average, respectively. With a sample preparation reduced to the minimum, characterization of the most abundant histones can be achieved in a single experiment. Semi-quantitative information can...