2.7. Enrichment of the acetylated peptides by immunoaffinity precipitation

To enrich lysine‐acetylated peptides, 3 μg TMT‐labeled peptides of each fraction and 1.5 mg label‐free peptides of each samples were dissolved in 300 μL NETN buffer (100 mM NaCl, 1 mM EDTA, 50 mM Tris‐HCl, 0.5% NP‐40, pH 8.0). The peptides were incubated with 20 μL prewashed antibody beads (cat no: PTM‐104, Jingjie PTM BioLabs, Hangzhou, China) at 4°C overnight with gentle shaking. The beads were then washed four times with NETN buffer and twice with ddH₂O. Next, the bound peptides were eluted from the beads using 0.1% trifluoroacetic acid. Then the eluted peptides were lyophilized using a lyophilizer. Finally, the resulting peptides were desalted with C18 ZipTips (Merck Millipore, USA) according to the manufacturer's instructions.