Stoichiometry of site-specific lysine acetylation in an entire proteome
Open accessAbstract
Acetylation of lysine ϵ-amino groups influences many cellular processes and has been mapped to thousands of sites across many organisms. Stoichiometric information of acetylation is essential to accurately interpret biological significance. Here, we developed and employed a novel method for directly quantifying stoichiometry of site-specific acetylation in the entire proteome of Escherichia coli. By coupling isotopic labeling and a novel pairing algorithm, our approach performs an in silico enrichment of acetyl peptides, circumventing the need for immunoenrichment. We investigated the function of the sole NAD(+)-dependent protein deacetylase, CobB, on both site-specific and global acetylation. We quantified 2206 peptides from 899 proteins and observed a wide distribution of acetyl stoichiometry, ranging from less than 1% up to 98%. Bioinformatic analysis revealed that metabolic enzymes, which either utilize or generate acetyl-CoA, and proteins involved in transcriptional and translational processes displayed the highest degree of acetylation. Loss of CobB led to increased global acetylation at low stoichiometry sites and induced site-specific changes at high stoichiometry sites, and biochemical analysis revealed altered acetyl-CoA metabolism. Thus, this study demonstrates that sirtuin deacetylase deficiency leads to both site-specific and global changes in protein acetylation stoichiometry, affecting central metabolism.
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Chemical labeling approaches, apart from the SILAC-based indirect method that overestimates due to cross-talk with other lysine modifications, indicate that average occupancy of most mitochondrial acetylation sites is below 1%.
“With the exception of the SILAC-based indirect stoichiometry method (which overestimates stoichiometry via crosstalk with other lysine modifications), all the chemical labeling methods have concluded that most of the mitochondrial protein acetylation sites are occupied at less than 1% on average.”
From The Mitochondrial Acylome Emerges: Proteomics, Regulation by Sirtuins, and Metabolic and Disease Implications (2018) · cites this paper as Baeza et al. 2014
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- Authors
- Baeza, J., Dowell, J. A., Smallegan, M. J., Fan, J., Amador-Noguez, D., Khan, Z., & Denu, J. M.
- Affiliations
- University of Wisconsin–Madison (US)
- Corresponding author
- Denu, John M.
- Venue
- Journal of Biological Chemistry
- Publisher
- Elsevier BV
- Published
- 2014-08
- Volume
- 289
- Issue
- 31
- Pages
- 21326-21338
- ISSN
- 0021-9258 (ISSN-L)
- PMC
- PMC4118097
- OpenAlex
- W2037555736
- Cited by
- 154
- References
- 41
- Citations by year
- 2026: 5 · 2025: 4 · 2024: 7 · 2023: 6 · 2022: 5 · 2021: 18 · 2020: 11 · 2019: 20 · 2018: 18 · 2017: 20 · 2016: 17 · 2015: 21 · 2014: 2
- FWCI
- 8.57
- Type
- journal article
- Language
- en
- Open access
- hybrid
- doi.org
- Licence
- Locations
- Journal of Biological Chemistry (publishedVersion, cc-by)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- Funders
- National Institutes of Health (GM065386)
- Topics
- Physical Sciences › Chemistry › Spectroscopy › Advanced Proteomics Techniques and Applications
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Ubiquitin and proteasome pathways
- Health Sciences › Medicine › Geriatrics and Gerontology › Sirtuins and Resveratrol in Medicine
- Keywords
- lysine acetylation, Escherichia coli proteome, Cobb, NAD+-dependent deacetylase, sirtuins, acetyl-CoA metabolism, isotopic labeling, metabolic enzymes
- MeSH terms
- Acetylation, Amino Acid Sequence, Bacterial Proteins, Chromatography, High Pressure Liquid, Escherichia coli, Lysine, Molecular Sequence Data, Computational Biology, Proteome, Tandem Mass Spectrometry
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