Table 2.
Kinetic Rate Constant Estimates determined by Surface Plasmon Resonance a
| Protein | kon (M−1s−1) | SD | koff (s−1) | SD | Kd (nM) | errorb | Ki (nM)c | Fold differencef |
|---|---|---|---|---|---|---|---|---|
| H1 WTd | 2.23×105 | 5.0×103 | 6.03×10−5 | 3.1×10−6 | 0.271 | 0.015 | 0.021 | 12.9 |
| H1 WTd,e | 4.85×105 | 2.06×105 | 7.10×10−5 | 5.6×10−6 | 0.146 | 0.063 | 0.021 | 7.0 |
| H2 WT | 1.28×105 | 1.54×104 | 2.76×10−4 | 2.32×10−5 | 2.16 | 0.32 | 2.11 | 1.02 |
| H2 WTe | 2.3×105 | 2.39×104 | 3.69×10−4 | 6.45×10−5 | 1.61 | 0.32 | 2.11 | 0.8 |
| H1 L59F | 1.12×105 | 7.00×103 | 0.2020 | 0.0351 | 2050 | 478 | 200 | 10 |
| H2 T125A | 7.21×104 | 4.61×103 | 5.3×10−3 | 3.0×10−4 | 73.5 | 6.3 | 53 | 1.4 |
| H2 T125Ae | 1.33×105 | 2.21×104 | 4.8×10−3 | 6.8×10−4 | 36.5 | 8.0 | 53 | 0.7 |
| H2 N127A | 3.48×104 | 6.81×102 | 0.0591 | 0.0084 | 1710 | 773 | 397 | 4.3 |
a
Microscopic rate constant estimates are the average of three individual sensorgrams.
b
The dissociation constant error was propagated from the on- and off-rate standard deviations.
c
The inhibition constants determined by progress curve analysis (WT) or 5×5 assay (EapH1 L59F, EapH2 T125A and N127A; Fig. 7).
d
Values previously determined by Herdendorf and Geisbrecht. 3.
e
Microscopic rate constant estimates determined by single-cycle binding kinetics.
f
The fold difference depicts the difference between the dissociation constant determined by SPR and the inhibition constant determined by kinetic analysis.