Table 4.
Characterisation of the CPAP:STIL interaction in vitro
| Danio rerio STIL404–448 peptide in syringe | Danio rerio CPAP937–1124 TCP domain in cell | Number of binding sites (N) | SD N | KD (μM) | SD KD (μM) | ΔH (kcal/mol) | SD (kcal/mol) | n (number of measurements) | Factor change in KD |
|---|---|---|---|---|---|---|---|---|---|
| WT | WT | 1.07 | 0.04 | 1.9 | 0.2 | −10.1 | 0.3 | 5 | 1 |
| WT | F978V | 0.70 | 0.09 | 37 | 10 | −23 | 5 | 4 | 20 |
| WT | T986V | 1.01 | 0.07 | 1.9 | 0.2 | −10.6 | 0.4 | 3 | 1 |
| WT | Y994V | 1.00 | 0.33 | 68 | 14 | −9.5 | 3.8 | 5 | 36 |
| WT | F1015V | 0.93 | 0.13 | 70 | 18 | −10.4 | 2.7 | 3 | 37 |
| WT | E1021V | 0.91 | 0.13 | 16 | 2 | −8.1 | 0.6 | 3 | 8 |
| WT | WT | 1.07 | 0.04 | 1.9 | 0.2 | −10.1 | 0.3 | 5 | 1 |
| P417A | WT | 1.06 | 0.02 | 37 | 1.3 | −11.5 | 0.2 | 3 | 20 |
| R418A | WT | 1.12 | 0.02 | 19 | 1 | −8.8 | 0.1 | 4 | 10 |
| P421A | WT | 1.16 | 0.03 | 17 | 0.3 | −9.8 | 0.2 | 4 | 9 |
| N422A | WT | 1.09 | 0.03 | 0.7 | 0.05 | −12.3 | 0.3 | 4 | 0.4 |
| P423A | WT | 1.16 | 0.05 | 4.6 | 0.3 | −10.9 | 0.4 | 4 | 2.4 |
Tables show the binding parameters between various D. rerio CPAP and STIL constructs obtained from ITC experiments. The measurements of the WT STIL404–448—WT CPAP937–1124 interaction are identical to each other and identical to those shown in Table 1 and are only presented again to allow easier comparison within each table. Fitting was performed with N as a variable. Constraining N to a fixed value of 1 during fitting produced KD values that were within the experimental error of those tabulated here. In control measurements on wild-type material and a selection of mutants of both CPAP and STIL, the experimental configuration was reversed with CPAP protein titrated into STIL peptide in the ITC cell. These experiments gave similar values for N, KD and ΔH to the standard configuration reported here.