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Assay Method Information |
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| | Inhibition Assay |
| Description: | Assays for slow-onset inhibitors were carried out by adding 1 nM PaMTIP into reaction mixtures at 25 °C. containing 100 mM Hepes, pH 7.4, 100 mM phosphate, pH 7.4, 2 mM MTI, 5 mM DTT, 0.5 unit of xanthine oxidase and variable inhibitor concentration. Inhibitors were present at >10 times the enzyme concentration. Assays for MTA inhibition used 200 μM MTI. Controls having no enzyme and no inhibitor were included in all of the inhibition assays. Slow-onset inhibitors display a second phase of tighter binding after reaching a thermodynamic equilibrium with the enzyme. The equilibrium constant for the second binding phase is indicated as Ki*. |
| Affinity data for this assay | |
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