Figure 6.

Chloroquine inhibits CTSL activity and increases intrinsic fluorescence in vitro. (A) CTSL hydrolysis of dipeptide substrate Z-FR-AMC was monitored by measuring the fluorescence of the released AMC. Reaction was performed at pH 5.5 in 100 mM MES-NaOH, 150 mM NaCl, and 7.5 mM DTT for 30 min after 25 min preincubation with chloroquine (CQ). Representative plot and calculation of three biological replicates. (B) Intrinsic fluorescence of purified CTSL-Flag, 200 nM, incubated with and without chloroquine. Representative data from three independent experiments. Fluorescence values were corrected for chloroquine background at these wavelengths. (C) Schematic representation of chloroquine-induced lysosome amino acid accumulation as compared with Bafilomycin treatment.