| Assay Method Information | |
| | Optical Titration Assay |
| Description: | Dissociation constants (Kd values) for binding of the substrates N-palmitoylglycine (NPG) and omeprazole (OMP) to WT and mutant BM3 heme domains were determined by UVvisible absorption titrations using ~1-4 μM protein in 100mMKPi, pH 7.0, at 25 °C (assay buffer) in 1-cm path length quartz cuvettes and as in our previous studies. Spectra were recorded for substrate-free enzymes and following addition of ligands during substrate titrations (typically 800 to 250 nm). Titrations were recorded until no further spectral changes were observed in the P450s. |
| Affinity data for this assay | |
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