FIGURE 6.

Determination of CatS activity in antigen presenting cells. Hydrolysis of the internally quenched fluorescent peptide PMGLP at a concentration of 20μm catalyzed by endosomal extracts from WT100 (A) and DCs (B). Endosomal fractions containing 2.75 μg (WT100) and 2.88 μg (DC) of total protein were preincubated for 1 h at 37 °C: ▪, without inhibitor; , with 10 nm LHVS. Product formation was recorded by fluorescence emission (λ_ex = 340 nm, λ_em = 405 nm). All of the assays were performed in triplicate at 37 °C in 50 mm sodium acetate buffer (pH 5.5). C, determination of endosomal CatS activity in antigen presenting cells. The initial velocities of the hydrolysis reactions were determined and referred to total protein amount in endosomal fractions. The heights of the bars denote endosomal CatS activity in μm min^–1 μg^–1. The results are representative of at least three independent experiments. The error bars indicate standard deviation.