Fig. 1.
Stimulation of RAW264 cells with LPS up-regulates and downregulates fatty-acylated proteins identified by metabolic labeling with 17ODYA and proteomic analysis. (A) Scheme of experimental procedure comprising metabolic labeling of cells with 50 μm 17ODYA or exposed to 0.05% DMSO carrier for 4 h, stimulation with 100 ng/ml LPS for 60 min, cell lysis, click chemistry reaction with biotin-azide, and capture of tagged proteins on streptavidin-coupled beads followed by mass spectrometry. Three experiments were performed. The abundance of proteins was estimated using a semiquantitative spectral counting approach described under “Experimental Procedures.” A total of 646 fatty-acylated proteins that were eventually identified with high and medium confidence are listed in supplemental Table 3. (B) Acylated proteins up- or downregulated or not affected by LPS treatment of cells revealed by calculating the RL ratio and listed in supplemental Table 4. Up-regulated and downregulated proteins include those (34 and 32 proteins listed in Table I) selected basing on the number of experiments they were detected in.