Table 1.
Kinetic parameters for TGR catalytic activities
| Substrates | TR1 | TGR | |
|---|---|---|---|
| Specific activity (μmol/min per mg) | NADPH + DTNB (TR activity) | 25 ± 3.0 | 3.1 ± 0.35 |
| NADPH + Trx (TR activity) | 5.9 ± 0.6 | 4.2 ± 0.6 | |
| NADPH + GSSG (GR activity) | ND | 2.0 ± 0.34 | |
| NADPH + HED (Grx activity) | ND | 1.9 ± 0.2 | |
| Apparent Km (μM) | DTNB | 212 | 14.7 |
| NADPH | 19.6 | 10.7 | |
| Trx | 4.73 | 3.0 | |
| GSSG | — | 8.84 | |
| HED | — | 45.2 | |
| Apparent kcat (min−1) | DTNB | 5,180 | 392 |
| Trx | 590 | 340 | |
| GSSG | — | 94 | |
| HED | — | 72 |
Apparent kinetic parameters were determined as described in Experimental Procedures using Lineweaver–Burk plots and varying concentrations of one substrate (2.5–500 μM GSSG, 10–500 μM HED, 0.069–0.69 μM Trx, or 1.25–425 μM DTNB) in a corresponding two-substrate reaction, while maintaining the concentration of a second substrate, NADPH, constant at 0.2 mM. Likewise, NADPH parameters were determined at 2–60 μM NADPH and 5 mM DTNB. ND, not detected.