| Assay Method Information | |
| | Binding Kinetics to KIF18a-Microtubule Complex |
| Description: | Table 10: Compound binding kinetics parameters (kon and koff) were determined by the method of global progress curve analysis (GPCA). KIF18A (0.25 nM) was incubated for up to 24 hr with serially diluted compound in the assay buffer containing 80 mM PIPES, pH 6.9, 1 mM ATP, 0.1 mg/ml preformed microtubule from porcine brain (Cytoskeleton), 1 mM MgCl2, 1 μM Taxol, 75 mM KCl, 1 mM EGTA, 1 mM DTT, 0.01% BSA and 0.005% Tween-20. ADP product levels were determined by the Promega ADP-Glo assay. The time/dose-dependent progress curves were then globally fit to a Michaelis-Menten kinetics model with 1-step slow binding inhibition to derive both on-rate kon and off-rate koff values. |
| Affinity data for this assay | |
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