Table 1.
Reaction | kdiss/s−1 | kass/M−1⋅s−1 | KD/nM |
---|---|---|---|
1. wtRBD + wtRas⋅mGppNHp | 7.4 | 4.5 × 107 | 160 |
2. RBD(ch) + wtRas⋅mGppNHp | 5.9 | 3.5 × 107 | 170 |
3. RBD/L91NBD + wtRas⋅GppNHp | 11 | 1.8 × 107 | 610 |
4. RBD/L91NBD + Ras⋅GppNHp(ch) | 11 | 2.3 × 107 | 480 |
5. wtRBD/L91W/H + wtRas⋅GppNHp* | 14 | 3.6 × 107 | 380 |
Reactions 1 and 2, kinetics were monitored by using the mant fluorescence of mGppNHp. Reactions 3 and 4, kinetics monitored by using the NBD-Dap fluorescence. Reaction 5, kinetics monitored by using the tryptophan fluorescence from ref. 7. wt, wild type; H, His tag.