1. Site-Specific Lysine Acetylation Stoichiometry Across Subcellular Compartments
- Author
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Anastasia J. Lindahl, John M. Denu, James A. Dowell, Alexis J. Lawton, and Josue Baeza
- Subjects
0303 health sciences ,Cell division ,Chemistry ,030302 biochemistry & molecular biology ,Lysine ,Mass spectrometry ,Proteomics ,Chromatin ,Cell biology ,03 medical and health sciences ,Acetylation ,Cell fractionation ,Fragmentation (cell biology) ,030304 developmental biology - Abstract
Posttranslational modifications of proteins control many complex biological processes, including genome expression, chromatin dynamics, metabolism, and cell division through a language of chemical modifications. Improvements in mass spectrometry-based proteomics have demonstrated protein acetylation is a widespread and dynamic modification in the cell; however, many questions remain on the regulation and downstream effects, and an assessment of the overall acetylation stoichiometry is needed. In this chapter, we describe the determination of acetylation stoichiometry using data-independent acquisition mass spectrometry to expand the number of acetylation sites quantified. However, the increased depth of data-independent acquisition is limited by the spectral library used to deconvolute fragmentation spectra. We describe a powerful approach of subcellular fractionation in conjunction with offline prefractionation to increase the depth of the spectral library. This deep interrogation of subcellular compartments provides essential insights into the compartment-specific regulation and downstream functions of protein acetylation.
- Published
- 2019
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