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Assay Method Information |
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| | Inhibition Assay |
| Description: | Inhibition constants (Ki, equivalent to Kd for competitive inhibitors) were measured at 25 °C by adding purified OPRT enzymes (25 nM for PfOPRT and 100 nM for HsOPRT) to reaction mixtures with 50mM Tris-HCl (pH 8.0), 5 mM MgCl2, 1 mM PPi, 0.5 mM OMP, and inhibitor at varied concentrations. In all cases, the inhibitor concentration was at least 10 times the enzyme concentration. Reaction rates were monitored through the absorbance increase at 295 nm to give an extinction coefficient of 3.67 mM-1 cm-1 [Abdo et al., Org. Lett., 12:2982-2985]. |
| Affinity data for this assay | |
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