Abstract
Top-down mass spectrometry is a valuable tool for understanding gene expression through characterization of combinatorial histone post-translational modifications (i.e., histone code). In this protocol, we describe a top-down workflow that employs liquid chromatography (LC) coupled to mass spectrometry (MS), for fast global profiling of changes in histone proteoforms, and apply LCMS top-down approach for comparative analysis of a wild-type and a mutant fungal species. The proteoforms exhibiting differential abundances can be subjected to further targeted studies by other MS or orthogonal (e.g., biochemical) assays. This method can be generally adapted for screening of changes in histone modifications between samples such as wild type vs. mutant or healthy vs. diseased.
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Acknowledgment
The authors thank Christopher Wilkins, Jung Kap Park, and Sangtae Kim at the Pacific Northwest National Laboratory (PNNL) for developing the bioinformatics software used in this work. We appreciate the help from other PNNL colleagues: Matthew Monroe and Nikola Tolić for data analysis and Rosalie K. Chu, Rui Zhao, Anil K. Shukla, and Ron Moore for running LCMS experiments. We also thank Jonathan Galazka at the Oregon State University for preparing the fungal histone samples. The research was performed in Environmental Molecular Sciences Laboratory (EMSL), a US Department of Energy (DOE) national user facility at the Pacific Northwest National Laboratory (PNNL) in Richland, WA.
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Zhou, M. et al. (2017). Profiling Changes in Histone Post-translational Modifications by Top-Down Mass Spectrometry. In: Wajapeyee, N., Gupta, R. (eds) Eukaryotic Transcriptional and Post-Transcriptional Gene Expression Regulation. Methods in Molecular Biology, vol 1507. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6518-2_12
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DOI: https://doi.org/10.1007/978-1-4939-6518-2_12
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