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| | PDK Inhibition Assay |
| Description: | To determine the IC50 for PDK inhibitors, a mixture containing 0.05-0.2 μM PDK, 6μM E1, with or without 0.5 μM of the PDC core E2/E3BP, and various amounts of inhibitor was incubated at 25 °C for 10 min in a buffer of 20 mM Tris-Cl, pH 7.5, 10 mM KCl, 5 mM MgCl2, 2 mM DTT, 0.02% (v/v) Tween 20, and 0.1 mg/ml bovine serumalbumin before the addition of 50 μM ATP to initiate the reaction. All inhibition titrations were performed at 10 dose points ranging from 31.6 to 1 mM in a 3.162-fold dilution series, with each inhibitor concentration tested in duplicate. The remaining steps were described previously [Wynn et al., J. Biol. Chem., 283:25305-25315]. |
| Affinity data for this assay | |
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