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What Is Protein acetylation?
Protein acetylation is a ubiquitous post-translational modification that occurs in both eukaryotic and prokaryotic organisms. It is the process of transferring and adding acetyl groups to protein lysine residues or protein N-terminal through either nonenzymatic or enzymatic means, with the latter one more common. It occurs irreversibly on the α-amino group at the N-terminal amino acid or reversibly on the ε-amino group on the side chain of the lysine residue. Most of the N-terminal acetylation modification occurs on eukaryotic, which is catalyzed by N acetyltransferase (NAT) and the acetylation modification on lysine is a reversible process, mainly catalyzed by lysine acetylase (KAT) and lysine deacetylase (KDAC). In contrast to reversible and dynamic Lys-acetylation, N-terminal acetylation modification is irreversible and persists throughout the protein's life.
Protein acylation have been shown to impact various cellular processes relevant to physiology and diseases, such as protein stability, protein subcellular localization, enzyme activity, transcriptional activity, protein–protein interactions and protein–DNA interactions. Importantly, dysfunction of this modification has been implied in many diseases, including aging, diabetes, cancer. Hence, the study of protein acetylation dynamics is critical for understanding of how this modification regulates protein stability, localization, and function.
Figure 1. Schematic outline of N-terminal and lysine protein acetylation.
The NTA is a highly abundant modification observed in 80%-90% of soluble proteins in human and plants, catalyzed by up to five ribosome-associated NAT complexes that typically occurring co-translationally. Several reports have shown that NAT is involved in various protein functions, such as subcellular targeting, protein interaction, folding, and turnover.
Many lysine residues can be acetylated, mainly on histone tails (sometimes in core). Lysine acetylation is not limited to histone proteins, as it has been reported to occur in non-histone proteins that play crucial roles in cellular metabolism, cell cycle regulation, aging, growth and development, angiogenesis, and cancer progression. Moreover, Lysine acetylated proteins appears in almost every compartment of a cell, such as nucleus, mitochondrion, and cytoplasm.
Our Acetyl-proteomics Service
Due to relatively low levels of acetylation modifications that mass spectrometric identification of acetylation sites is not trivial. Hence, immunoaffinity enrichment strategies in combination with high-resolution mass spectrometry have proven to be highly effective to gain deeper insights into the dynamic acetylome.
Classically, the workflow of our acetyl-proteomics includes protein extraction, one or multiple enzymes for full proteolytic digestion into peptides, immunoaffinity enrichment of acetylated peptides, identification of acetylation sites, and relative quantification of changes for identified acetylation sites. Additionally, the application of MS analysis can be complemented by incorporating SILAC or iTRAQ/TMT labeling techniques to enhance the precision of relative quantitation between samples.
Figure 2. General workflow elements for Acetylome characterization in biological samples. Abbreviation: Ac-Acetylation.
- Professional detection and analysis capability: Experienced PTM research team, strict quality control system, together with ultra-high resolution detection system and professional data pre-processing and analysis capability, ensure reliable and accurate data.
- Reproducible: Obtain consistent and reproducible inter- and intra- assay results for data analysis.
- High specificity: Use acetylation-specific antibodies for acetyl-peptides enrichment.
- Multiplex, high-throughput: Deeper Coverage of Acetylation Site Identification.
- High resolution and sensitivity: Q-Exactive, Q-Exactive HF, Orbitrap Fusion™ Tribrid™.
Tissue: animal tissue > 50 mg;
fresh plant > 100 mg;
Cell: suspension cell > 2 x 107;
adherent cell > 2 x 107;
microorganism > 50 mg or 2 x 107 cells;
Body fluid: Serum/plasma > 500 μL;
Protein: Total protein >1 mg and concentration >1 μg/μL.
Note: In order to ensure the test results, please inform the buffer components if you give us proteins, whether it contains thiourea, SDS, or strong ion salts. In addition, the sample should not contain components such as nucleic acids, lipids, and polysaccharides, which will affect the separation effect.
- Detailed report, including experiment procedures, parameters, etc.
- Raw data and data analysis results
How to place an order:
At Creative Proteomics, many excellent and experienced experts will optimize the experimental protocol according to your requirement and guarantee the high-quality results for protein acetylation. Creative Proteomics provides a broad range of technologies for acetylation research that enable quantification of protein amount and acetylation modification. Please feel free to contact us by email to discuss your specific needs. Our customer service representatives are available 24 hours a day, from Monday to Sunday.
- Gibbs DJ, Bailey M, Etherington RD. A stable start: cotranslational Nt-acetylation promotes proteome stability across kingdoms. Trends in Cell Biology. 2022 May;32(5):374-376.
- Aksnes H, Ree R, Arnesen T. Co-translational, Post-translational, and Non-catalytic Roles of N-Terminal Acetyltransferases. Molecular Cell. 2019 Mar 21;73(6):1097-1114.
- Shang S, Liu J, Hua F. Protein acylation: mechanisms, biological functions and therapeutic targets. Signal Transduction and Targeted Therapy. 2022 Dec 29;7(1):396.
In-depth Profiling and Quantification of the Lysine Acetylome in Hepatocellular Carcinoma with a Trapped Ion Mobility Mass Spectrometer
Journal: Molecular & Cellular Proteomics
Main Technology: Label-free quantitative proteomics
Hepatocellular carcinoma (HCC) is the third most common cause of cancer-related death worldwide with limited therapeutic options. Comprehensive investigation of protein posttranslational modifications in HCC is still limited. Lysine acetylation is one of the most common types of posttranslational modification involved in many cellular processes and plays crucial roles in the regulation of cancer. With the advance in modern mass spectrometry, more and more K-acetylation sites are being characterized, however, how majority of these sites are regulated and more importantly how they are involved in diseases remain largely unclear.
The identification results of Lysine Acetylome
We analyzed the proteome and K-acetylome in eight pairs of HCC tumors and normal adjacent tissues using a timsTOF Pro instrument. As a result, we identified 9219 K-acetylation sites in 2625 proteins, of which 1003 sites exhibited differential acetylation levels between tumors and normal adjacent tissues. Importantly, many novel tumor-specific K-acetylation sites were characterized. We observed an overall suppression of protein K-acetylation in HCC tumors, especially for enzymes from various metabolic pathways. We found the expression of deacetylase sirtuin 2 (SIRT2) was upregulated in HCC tumors and overexpression of SIRT2 in HCC cells inhibited both glycolysis and oxidative phosphorylation. Our findings provide valuable information to better understand the roles of K-acetylation in HCC and to treat this disease by correcting the aberrant acetylation patterns.
Figure 1. Workflow of quantitative acetylome in HCC.