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| | ATPase Assay |
| Description: | The conversion of ATP to ADP by S. aureus TopoIV enzyme is coupled to the conversion of NADH to NAD+, and the progress of the reaction is measured by the change in absorbance at 340 nm. TopoIV (64 nM) is incubated with the selected compound (3% DMSO final) in buffer for 10 minutes at 30 ° C. The buffer consists of 100 mM Tris 7.5, 1.5 mM MgCl2, 200 mM K.Glutamate, 2.5 mM phosphoenol pyruvate, 0.2 mM NADH, 1 mM DTT, 5 g/mL linearized DNA, 50 g/mL BSA, 30 g/mL pyruvate kinase, and 10 g/mL lactate dehyrodgenase (LDH). The reaction is initiated with ATP, and rates are monitored continuously for 20 minutes at 30 ° C. on a Molecular Devices SpectraMAX plate reader. |
| Affinity data for this assay | |
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