Figure 6: Specificity of fluorogenic peptides in complex proteomes.

a) Activity of endogenous depalmitoylases measured in T. gondii lysates (at 5 μg) using the optimized fluorgenic substrates (at 2.5 μM). Error bars represent the S.D. of three replicates. b) Normalized rate of hydrolysis for the ASC₂₀KRNT substrate (at 2.5 μM) measured in Ku80 T. gondii lysates (at 5 μg) untreated (WT) or treated with JCP174 (JCP174, 10 μM) or Palmostatin B (PalmB, 10 μM) and untreated PPT1 knockout (ΔPPT1) T. gondii lysates (at 5 μg). Error bars represent S.D. of three biological replicates and 3 technical replicates. Statistical significance is calculated using one-way ANOVA (**** p < 0.0001), (*** p = 0.0003), (* p = 0.0252). c) Activity of endogenous depalmitoylases measured in mammary breast tumor homogenate (at 5 μg) using optimized and non-optimal fluorogenic substrates (at 2.5 μM). Error bars represent the S.D. of three replicates. d) and e) Normalized rate of hydrolysis for ASC₂₀KKNT and AHC₂₀DRNT (at 2.5 μM) measured in tumor homogenates treated with vehicle (DMSO) or the inhibitors ML348, ML349 and Palmostatin B (at 10 μM). Error bars represent S.D. of three biological replicates and 3 technical replicates. Statistical significance is calculated using one-way ANOVA (**** p = 0.0001), (ns p = 0.6711). See also figure S5.