| Assay Method Information | |
| | In vitro Enzyme Inhibition Assay |
| Description: | The assays were conducted at 37 °C in a mixture containing 100 mM Tris/Cl pH 7.9, 50 mM KCl, 16 mM MgCl2, 5 mM ATP, 3 mM DTT, 4 mg/ml E. coli MRE600 tRNA (Roche) and 10 μM L-tyrosine (0.3 μM L-[ring-3,5-3H] tyrosine (PerkinElmer, Specific activity: 1.48-2.22 TBq/mmol), 10 μM carrier). Above all, TyrRS (0.2 nM) was pre-incubated with kinds of concentrations inhibitor for 10 min (RT) followed by the addition of pre-warmed mixture and moved them together to 37 °C. After specific intervals, the reaction was ended by the addition of aliquots of the reaction mix into gelid 7% trichloroacetic acid and harvested onto 0.45 mm hydrophilic Durapore filters (Millipore Multiscreen 96-well plates) and counted by liquid scintillation. |
| Affinity data for this assay | |
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