Selenosulfide inhibitors of STEP. (a) Combining the STEP-specific
aryl scaffold of previously developed active site directed reversible
inhibitors 5 and 6 with the selenosulfide
PTP pharmacophore to generate potent STEP selenosulfide inhibitor 7 assayed with or without physiological intracellular concentrations
of GSH (1 mM). (b) Irreversible inhibition with 7. STEP
(200 nM) and inhibitor 7 (1 μM) (or DMSO control)
were incubated for 60 min in the absence of GSH to inhibit enzymatic
activity prior to dialysis. The enzymatic activity of STEP was measured
using the pNPP assay at 0, 5, and 24 h after dialysis. (c) Reactivation
of inhibited STEP by the addition of thiols. STEP (200 nM) and 7 (5 μM) were incubated for 10 min, and following addition
of GSH or DTT (1 mM each) or water (no reductant), for another 0,
0.5, 1, and 1.5 h after which enzymatic activity was measured. (d)
Structure–activity relationship of STEP selenosulfide inhibitors.