Dynamics of Post-Translational Modifications on Human Histone H4 Through the Cell Cycle

摘要

Detection and characterization of histone PTMs have been conducted by antibodies, Bottom Up MS, and Top Down MS. As analytical techniques, however, they are not germane to quantitative biology unless they can provide consistently reproducible data that can be used to extrapolate trends and make correlations in addition to answering questions regarding modifications singly and in combination. Hydrophilic interaction liquid chromatography (HILIC) is a 2D-LC fractionation technique that produces an acetylation-methylation dependent elution profile of oxidized H4 thereby allowing for more in-depth interrogation of the protein forms present in solution. The substrates for Top Down Mass Spectrometry (TDMS) are whole proteins, facilitating analysis of posttranslational modifications singly and in combination. Here we show that the combination of HILIC and TDMS enables reproducible quantitation of histone H4 PTMs as the cell cyles with the ability to observe biological phenomena such as progressive methylation and to detect rare modified protein forms.