2026-10-14T23:46:05Z https://www.scidb.cn/oai

10.57760/sciencedb.j00139.00299 2026-05-14T11:08:47Z

2026-05-14 Palmitoyl-proteomic profiling of large yellow croaker monocytes/macrophages with or without zDHHC20b knockdown during Pseudomonas plecoglossicida infection doi:10.57760/sciencedb.j00139.00299 en RAW mass spectrometry proteomics data of precipitated beads from the ABE assay of scrambled siRNA or zDHHC20b siRNA transfected and Pseudomonas plecoglossicida infected large yellow croaker MO/MΦ cells were subjected to on-bead tryptic digestion. Trypsin was first added to a final concentration of 20 ng/μL, and samples were digested overnight. Disulfide bonds were then reduced with 5 mM dithiothreitol (DTT) at 37°C for 60 min, followed by alkylation with 11 mM iodoacetamide (IAA) at room temperature in the dark for 45 min. Trypsin was subsequently added twice at a final concentration of 10 ng/μL, with digestion continued for 4 h each time. The resulting peptides were acidified and desalted. Peptides were dissolved in solvent A (0.1% formic acid, 2% acetonitrile in water) and loaded onto a homemade reversed-phase analytical column (25 cm×100 μm i.d.). Separation was performed on a NanoElute UHPLC system (Bruker Daltonics) at a flow rate of 500 nL/min using the following gradient of solvent B (0.1% formic acid in acetonitrile): 0-14 min, 6-24% B; 14-16 min, 24-35% B; 16-18 min, 35-80% B; 18-20 min, 80% B. Peptides were analyzed on a timsTOF Pro mass spectrometer (Bruker Daltonics, USA) in dia-PASEF mode with a capillary electrospray voltage of 1.75 kV. Full MS scans were acquired over m/z 300-1500, with 20 PASEF-MS/MS scans per cycle. The MS/MS scan range was m/z 400-8500, and the isolation window was 7 m/z. Raw DIA data were processed using DIA-NN (v1.8). Trypsin/P was specified as the cleavage enzyme with up to one missed cleavage allowed. N-terminal methionine excision and carbamidomethylation of cysteine were set as fixed modifications. The false discovery rate (FDR) was controlled at <1%. Functional enrichment analysis of differentially palmitoylated proteins was performed using Gene Ontology (GO) and KEGG pathway annotations. GO IDs were assigned using eggNOG-mapper, and proteins were classified into biological process, cellular component, and molecular function categories. KEGG annotation was performed by BLAST search against the KEGG database (e-value ≤1e-4). Fisher's exact test identified significantly enriched terms, defined as those with fold enrichment >1.5 and p-value <0.05. The selection criteria for differentially palmitoylated proteins were defined as an absolute log2 (fold change) ≥1 (corresponding to a ≥2-fold alteration in protein abundance), in conjunction with a false discovery rate (FDR)-adjusted p-value <0.05 as the significance threshold.  Palmitoylation; zDHHC20b; large yellow croaker; Mass spectrometry Zi-Yue Zhao Chenjie Fei NIE LI Jiong Chen PUBLIC https://creativecommons.org/licenses/by-nc-sa/4.0/ dataset http://www.doi.org/10.24272/j.issn.2095-8137.2025.573 Science Data Bank