Mechanism of Quantitative Acetylomics

Acetylation is a crucial post-translational modification (PTM) that plays a key role in various biological processes by regulating protein activity, function, and interactions. Quantitative acetylomics aims to systematically analyze the dynamic changes of protein acetylation and reveal their functional roles in physiological and pathological conditions. In recent years, with the advancement of mass spectrometry technology, researchers have been able to conduct high-throughput quantitative analysis of acetylated proteins in cells and tissues. This has opened unprecedented opportunities to explore the mechanisms of acetylation and its role in diseases.

 

Biological Mechanisms of Acetylation

Acetylation mainly occurs on lysine (Lys) residues and is catalyzed by acetyltransferases (KATs), which transfer an acetyl group from acetyl-CoA to the ε-amino group of lysine residues. This process occurs in the nucleus, mitochondria, and cytoplasm. Conversely, deacetylases (KDACs) can remove the acetyl group from lysine, thereby reversing the acetylation modification. The balance between KATs and KDACs is critical for regulating processes such as cell proliferation, metabolism, and apoptosis.

 

Acetylation modifies protein function, often by altering protein structure, stability, and interactions with other molecules. For example, histone acetylation can regulate chromatin structure and thereby influence gene expression. Additionally, acetylation can regulate non-histone proteins, such as metabolic enzymes, transcription factors, and signaling proteins, playing roles in processes like metabolism, inflammation, and cancer.

 

Quantitative Acetylomics Techniques

In quantitative acetylomics, commonly used analytical methods include liquid chromatography-mass spectrometry (LC-MS/MS) and isotope labeling techniques such as SILAC and TMT. These techniques allow researchers to quantitatively analyze the expression levels of acetylated proteins and the abundance of acetylation sites under different conditions. Below are two key techniques:

 

1. SILAC (Stable Isotope Labeling with Amino Acids in Cell Culture)

This method involves incorporating heavy isotope-labeled amino acids into the cell culture system to label acetylated proteins, followed by comparative analysis using mass spectrometry. SILAC offers precise labeling, ease of operation, and efficient quantitative analysis.

 

2. TMT (Tandem Mass Tags)

This method uses peptides labeled with different mass tags, allowing samples to be distinguished during mass spectrometry analysis. TMT enables high-throughput, multi-sample quantitative analysis, making it particularly suitable for complex samples.

 

Quantitative Analysis in Acetylomics

Mass spectrometry plays a crucial role in quantitative acetylomics analysis. First, samples undergo protein extraction and enzymatic digestion to generate peptides. Using specific enrichment techniques (such as anti-acetyl-lysine antibody enrichment), acetylated peptides are isolated and analyzed by LC-MS/MS. Mass spectrometry data provide information about acetylation sites, and in combination with isotope labeling or tagging techniques, researchers can quantify changes in acetylation under different conditions.

 

Quantitative results in acetylomics often require bioinformatics analysis for interpretation. For example, by comparing the changes in acetylation levels between experimental and control groups, researchers can identify acetylated proteins that play key roles in specific biological processes. Furthermore, pathway analysis and network analysis can reveal the regulatory mechanisms of acetylation in signaling pathways.

 

Research Progress in Acetylation Mechanisms

Recent studies have shown that acetylation is not limited to histone modifications in the nucleus but is also widespread in non-histone proteins, such as metabolic enzymes in the cytoplasm and mitochondria. For instance, in glycolysis, acetylation can regulate the activity of key enzymes, thereby affecting cellular energy metabolism. Moreover, acetylation is closely associated with various diseases, particularly cancer, neurodegenerative diseases, and metabolic disorders, and its mechanisms are increasingly being recognized.

 

Through quantitative acetylomics research, scientists can gain deeper insights into the dynamic regulatory mechanisms of acetylation and its functions in diseases, providing important theoretical foundations for developing new therapeutic strategies.