Table of Contents
International Journal of Proteomics
Volume 2014, Article ID 730725, 8 pages
Research Article

A Method to Determine Lysine Acetylation Stoichiometries

1Biological Science Division and Environmental, Pacific Northwest National Laboratory, Richland, WA 99352, USA
2Bindley Bioscience Center, Discovery Park, Purdue University, West Lafayette, IN 47907, USA
3Molecular Sciences Laboratory, Pacific Northwest National Laboratory, Richland, WA 99352, USA
4Institute for Basic Science, Seoul National University, Seoul 151-747, Republic of Korea

Received 13 March 2014; Accepted 11 June 2014; Published 20 July 2014

Academic Editor: Andrew J. Link

Copyright © 2014 Ernesto S. Nakayasu et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


Lysine acetylation is a common protein posttranslational modification that regulates a variety of biological processes. A major bottleneck to fully understanding the functional aspects of lysine acetylation is the difficulty in measuring the proportion of lysine residues that are acetylated. Here we describe a mass spectrometry method using a combination of isotope labeling and detection of a diagnostic fragment ion to determine the stoichiometry of protein lysine acetylation. Using this technique, we determined the modification occupancy for ~750 acetylated peptides from mammalian cell lysates. Furthermore, the acetylation on N-terminal tail of histone H4 was cross-validated by treating cells with sodium butyrate, a potent deacetylase inhibitor, and comparing changes in stoichiometry levels measured by our method with immunoblotting measurements. Of note we observe that acetylation stoichiometry is high in nuclear proteins, but very low in mitochondrial and cytosolic proteins. In summary, our method opens new opportunities to study in detail the relationship of lysine acetylation levels of proteins with their biological functions.