TABLE 1.
Kinetic parameters for binding of FVIIIa subunits and FIXa
Kinetic parameters were determined by SPR as described under “Experimental Procedures” using FVIII or FVIII(a)-derived subunits in the mobile phase and bFPR-FIXa bound to a SA chip. Samples were run at least three times, and mean values with standard deviations shown. Values for Kd and χ2 are shown for both the two-state and single-site models for comparison.
| SA analyte | ka1 | kd1 | ka2 | kd2 | Two-state |
Single-site |
||
|---|---|---|---|---|---|---|---|---|
| Kd | χ2 | Kd | χ2 | |||||
| m−1 s−1 | s−1 | s−1 | s−1 | nm | nm | |||
| A2 | (2.3 ± 0.7) × 107 | (0.52 ± 0.2) | (1.3 ± 0.2) × 10−2 | (2.6 ± 0.8) × 10−3 | 3.9 ± 1.6 | 0.87 | 3.9 ± 0.4 | 2.54 |
| A3C1C2 | (4.1 ± 0.8) × 105 | (0.9 ± 0.4) × 10−2 | (3.5 ± 0.6) × 10−3 | (0.7 ± 0.1) × 10−3 | 3.6 ± 0.6 | 135 | 11.3 ± 0.1 | 101 |
| a3A3C1C2 | (6.6 ± 0.7) × 105 | (3.0 ± 0.2) × 10−2 | (8.2 ± 1.4) × 10−3 | (2.4 ± 0.6) × 10−3 | 10.5 ± 0.9 | 6.6 | 16.7 ± 0.2 | 115 |
| A1/A3C1C2 | (6.6 ± 0.4) × 105 | (1.7 ± 0.2) × 10−2 | (7.0 ± 1.6) × 10−3 | (3.1 ± 1.1) × 10−3 | 7.5 ± 0.9 | 15 | 12.7 ± 0.3 | 90.0 |
| FVIII | (8.0 ± 1.0) × 105 | (3.1 ± 0.2) × 10−2 | (8.8 ± 0.3) × 10−3 | (2.1 ± 0.1) × 10−3 | 7.3 ± 0.4 | 6.7 | 11.1 ± 0.8 | 238 |