Table 1.
Steady-state kinetic parameters for AncFMO1
| Substrate | Fixed substrate | kcat (s−1) | KM (μM) | kcat/KM (s−1/M) |
|---|---|---|---|---|
| aBenzydamine | 0.10-mM NADPH | 0.18 (0.004) | <13 | >14,000 |
| aThioanisole | 0.10-mM NADPH | 0.19 (0.007) | <25 | >7600 |
| bNADPH | 1.0-mM thioanisole | 0.26 (0.009) | <9 | >29,000 |
| bNADH | 1.0-mM thioanisole | 0.53 (0.03) | 54 (8) | 10,000 |
| cNADPH | - | 0.13 (0.01) | 55 (13) | 2400 |
| cNADH | - | 0.11 (0.009) | 91 (17) | 1200 |
AncFMO, ancestral flavin-containing monooxygenase.
a
Reactions were followed by measuring the NADPH consumption at 340 nm and rates calculated using NADPH extinction coefficient 6.22 mM−1 cm−1.
b
NAD(P)H steady-state kinetics in the presence of 1.0-mM thioanisole, rates followed at 340 nm.
c
AncFMO1 uncoupling reaction with NAD(P)H, in the absence of the substrate. The values in brackets refer to SDs.