Parameters for FVa2 (FV(657-709)) and FVa3 (FV(1481-1545)) binding to immobilized α-thrombin, measured by SPR
| Analyte . | Chip* . | Binding model† . | ka (M−1s−1) . | kd (s−1) . | KD, μM‡ . |
|---|---|---|---|---|---|
| FV(695-699) | CM5-FPR-T | No binding | — | — | — |
| FV(695-701) | CM5-FPR-T | No binding | — | — | — |
| FV(657-679) | CM5-FPR-T | No binding | — | — | — |
| FV(679-701) | CM5-FPR-T | No binding | — | — | — |
| FV(657-709) | CM5-FPR-T | 1:1 | 3.69 × 103 (± 49.4) | 7.60 × 10−3 (± 1.47 × 10−4) | 2.06 ± 0.05 |
| FV(657-709) | SA-FPR-T | 1:1 | 1.33 × 103 (± 14.4) | 2.38 × 10−3 (± 2.09 × 10−5) | 1.95 ± 0.03 |
| FV(1481-1545) | CM5-FPR-T | 1:1 | 0.999 × 103 (± 8.42) | 1.94 × 10−3 (± 2.98 × 10−5) | 1.96 ± 0.03 |
| FV(1481-1545) | SA-FPR-T | 1:1 | 5.46 × 103 (± 65.3) | 1.60 × 10−3 (± 3.65 × 10−5) | 0.237 ± 0.006 |
| Analyte . | Chip* . | Binding model† . | ka (M−1s−1) . | kd (s−1) . | KD, μM‡ . |
|---|---|---|---|---|---|
| FV(695-699) | CM5-FPR-T | No binding | — | — | — |
| FV(695-701) | CM5-FPR-T | No binding | — | — | — |
| FV(657-679) | CM5-FPR-T | No binding | — | — | — |
| FV(679-701) | CM5-FPR-T | No binding | — | — | — |
| FV(657-709) | CM5-FPR-T | 1:1 | 3.69 × 103 (± 49.4) | 7.60 × 10−3 (± 1.47 × 10−4) | 2.06 ± 0.05 |
| FV(657-709) | SA-FPR-T | 1:1 | 1.33 × 103 (± 14.4) | 2.38 × 10−3 (± 2.09 × 10−5) | 1.95 ± 0.03 |
| FV(1481-1545) | CM5-FPR-T | 1:1 | 0.999 × 103 (± 8.42) | 1.94 × 10−3 (± 2.98 × 10−5) | 1.96 ± 0.03 |
| FV(1481-1545) | SA-FPR-T | 1:1 | 5.46 × 103 (± 65.3) | 1.60 × 10−3 (± 3.65 × 10−5) | 0.237 ± 0.006 |
— indicates not applicable; FV, factor V; and SPR, surface plasmon resonance.
CM5-FPR-T, FPR-inhibited thrombin immobilized on CM5 chips, most likely via exosite II lysine residues; SA-FPR-T, biotin-FPR–inhibited thrombin immobilized on SA chips; both exosites are presumably accessible.
Binding data was fitted using the BIAevaluation 4.1 software. The binding model chosen represents that with the lowest χ2 value.
Error propagated from experimental errors of ka/kd values.