Table 1.
Comparison of simulation results and experimental kinetic parameters for the wild-type or mutated off7 interacting with MBP
| Sample | Robetta simulation |
Single-molecule force microscopy |
Surface plasmon resonance |
|||
|---|---|---|---|---|---|---|
| ΔΔGbind (kcal/mol) | k0off (×10−3 s−1) | xβ (nm) | kon (×105 M−1 s−1) | koff (×10−3 s−1) | Kd (nM) | |
| Wild-type | — | 6.1 ± 2 | 0.59 ± 0.03 | 9.41 ± 0.4 | 7.3 ± 0.5 | 7.7 ± 0.6 |
| Y125H | 0.41 ± 0.03 | 47.3 ± 9 | 0.54 ± 0.02 | 6.14 ± 0.7 | 25.9 ± 0.8 | 42.2 ± 4.6 |
| D110A | 0.61 ± 0.10 | 262.8 ± 58 | 0.43 ± 0.03 | 2.45 ± 0.2 | 28.6 ± 1.2 | 116.7 ± 10.7 |
| Y81A | 1.88 ± 0.09 | N.B. | — | 0.64 ± 0.1 | 103.3 ± 7.8 | 1614 ± 280 |
| F79A | 2.79 ± 0.16 | N.B. | — | N.B. | — | — |
| W90A | 2.74 ± 0.19 | N.B. | — | N.B. | — | — |
| E3_5 | — | N.B. | — | N.B. | — | — |
N.B. represents no binding that is defined as <5 RU response was detected in SPR assay up to 1 μM concentration or <5% of successful binding events observed in single-molecule force microscopy.