Table IV.
Binding characteristics and metabolism of the alternate ligands of CYP73A1
Measurements were performed at 27°C with recombinant yeast microsomes. Dissociation constant for the enzyme-ligand complex and spin change were calculated from the saturation curves of difference spectra recorded upon ligand binding to the oxidized microsomes. Activities were measured at saturating substrate concentrations (150 and 400 μm). Extracted products were analyzed and quantified by HPLC and diode array detection. Activities were related to that measured with cinnamate as substate: 54 pkat.mg-1 protein. Inhibition of C4H activity was competitive. n.d. refers to not determined.
| Ligand | Binding Characteristics | Relative Activity | KI | |
|---|---|---|---|---|
| KD | Spin Change | |||
| CA | μM | % | % | μM |
| 8.8 ± 0.3 | 100 ± 1.4 | 100 ± 0.6 | - | |
| 3TA | 7.7 ± 0.6 | 101 ± 0.8 | 29 ± 0.7 | 2.1 ± 0.1 |
| 2TA | 6.3 ± 0.4 | 100 ± 1.5 | 41 ± 1.1 | 2.3 ± 0.2 |
| 4VB | 12 ± 0.9 | 87 ± 2.5 | 8.1 ± 0.4 | 5.6 ± 0.7 |
| 4FC | 9 ± 0.9 | 57 ± 1.7 | no metabolism | n.d. |