Analysis of Histones and Histones-Associated Proteins Using Mass Spectrometric Techniques
Nicklay, Joshua James, Department of Chemistry, University of Virginia
Hunt, Donald F., Department of Chemistry, University of Virginia
The epigenetic regulation of gene expression is directly linked to the structure that DNA forms within the nucleus of a eukaryotic cell. DNA is tightly compacted into chromatin together with small proteins known as histones. The posttranslational modifications of these histone proteins are known to be critical regulators of the transcriptional activity of the genome. Mass spectrometry has become a powerful technique for the characterization of proteins and for the identification of post-translational modifications. Described in this dissertation are two projects that utilize high resolution mass spectrometry in combination with collision activated dissociation (CAD) and electron transfer dissociation (ETD) tandem mass spectrometry for the characterization of posttranslational modifications on two groups of chromatin-associated proteins. In the first project, the combinations of modifications along the N-terminal tail domains of histone H3 and H4 were analyzed from five unique, Xenopus laevis cell types in order to profile the developmental changes that occur in the epigenetic regulation of gene expression throughout early embryogenesis. In the second project, the changes in the modification states of the histone chaperone protein nucleoplasmin were analyzed between two unique, oogenic states in order to fully characterize its role in the early embryogenesis and to fully elucidate the mechanisms of activation of its histone-chaperoning ability.
PHD (Doctor of Philosophy)
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