Figure 4.

HSP90 inhibitors inhibit NS2/3 cleavage in a cell-based assay. Cloned Jurkat cells expressing a fusion protein of NS2/3 in which BLA activity is the indicator of successful NS2/3 cleavage were treated with either geldanamycin or radicicol (see Materials and Methods). Inhibitor treatment was for 5 h, followed by addition of cycloheximide to stop protein synthesis (30 min) and subsequent addition of the BLA substrate, CCF2 (20). After 2 h, BLA activity was quantified by fluorescence readings (460-nm/530-nm ratio). Lower 460-nm/530-nm ratios indicate inhibition of NS2/3 cleavage. The geldanamycin (●) IC₅₀ is 40 nM and the radicicol (▵) IC₅₀ is 13 nM.