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| | Enzyme Inhibition and Ki Determination |
| Description: | The IC50 inhibition assays were performed in a 96-well microtiter plate format using purified recombinant IDO, which was added to the substrate, L-tryptophan (L-Trp), and the inhibitor. After the reaction, the product N-formylkynurenine was converted to kynurenine. The yellow color generated from reaction with kynurenine was measured at 490 nm using a microtiter plate reader. The data were analyzed using Graph Pad Prism 4 software (Graph Pad Software Inc.). For the Ki determinations, tryptophan concentrations were varied from 25 to 200 uM (Km= 42 uM) and inhibitor concentrations were varied between 3-fold above and below the calculated IC50. Data were analyzed with the Enzyme Kinetics module in SigmaPlot, version 10. |
| Affinity data for this assay | |
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