TABLE 1.
Thermodynamic analysis by ITC of interactions between ANAC013 wild-type and mutant fragments and RCD1-RST(499–572)
All experiments were performed as described under “Experimental Procedures.” RCD1 was in the syringe, and the ANAC013 peptides were in the cell. The standard errors for ΔH, Kd, and N were obtained from Origin when fitting the data to a one set of sites binding model.
| Protein | Kd | N | ΔH | −TΔS | ΔG |
|---|---|---|---|---|---|
| nm | kJ/mol | kJ/mol | kJ/mol | ||
| ANAC013(161–498)a | 537 ± 105 | 0.96 ± 0.03 | −41.0 ± 2.0 | 5.4 | −35.6 |
| ANAC013(232–299) | 92 ± 13 | 0.90 ± 0.02 | −51.9 ± 1.3 | 11.7 | −40.3 |
| ANAC013(254–299) | 32 ± 12 | 0.80 ± 0.01 | −27.2 ± 0.6 | −15.6 | −42.6 |
| ANAC013(254–274) | 9 ± 4 | 0.80 ± 0.01 | −45.0 ± 0.8 | −0.6 | −45.6 |
| ANAC013(254–268) | 595 ± 117 | 1.02 ± 0.04 | −27.6 ± 1.4 | −7.9 | −35.5 |
| ANAC013(254–274; E256A) | 43 ± 24 | 0.94 ± 0.02 | −25.5 ± 1.2 | −16.5 | −42.0 |
| ANAC013(254–274; D258A) | 61 ± 21 | 0.92 ± 0.02 | −32.8 ± 1.0 | −8.4 | −41.2 |
| ANAC013(254–274; D258P) | 110 ± 35 | 0.78 ± 0.02 | −33.4 ± 1.0 | −6.3 | −40.1 |
| ANAC013(254–274; M259A) | 98 ± 45 | 0.86 ± 0.04 | −26.8 ± 1.6 | −13.2 | −40.0 |
| ANAC013(254–274; Y260A) | 1114 ± 346 | 1.05 ± 0.07 | −16.5 ± 1.6 | −17.5 | −34.0 |
| ANAC013(254–274; L261A) | 885 ± 272 | 1.04 ± 0.07 | −13.6 ± 1.2 | −20.9 | −34.5 |
| ANAC013(254–274; E262A) | 436 ± 100 | 0.94 ± 0.03 | −30.7 ± 1.5 | −5.5 | −36.2 |
| ANAC013(254–274; I263A) | 43 ± 22 | 0.83 ± 0.02 | −35.2 ± 1.3 | −6.8 | −42.0 |
| ANAC013(254–274; N264A) | 64 ± 10 | 0.74 ± 0.01 | −34.5 ± 0.5 | −6.6 | −41.1 |
| ANAC013(254–274; N264K) | NBb | ||||
| ANAC013(254–274; D265A) | NB | ||||
| ANAC013(254–274; D265N) | NB | ||||
| ANAC013(254–274; L266A) | NB | ||||
| ANAC013(254–274; M267A) | 97 ± 26 | 0.87 ± 0.02 | −35.4 ± 1.5 | −4.6 | −40.0 |
a Data were determined as described by O'Shea et al. (16).
b NB means no detectable binding.