Appendix 1—table 2. ITC analysis of all Cd2+-binding isotherms, with chosen model and values shaded.
Appendix 1—table 2—source data 1. Source files (Origin files) of ITC experiments of cadmium binding to each DraNramp construct.
elife-84006-app1-table2-data1.zip (3.7MB, zip)
| Protein construct | Two-site sequential binding model (fixed n=2) | One-site fit with fixed n=1 | ||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Kd1(µM) | ΔH1(kcal/mol) | -TΔS1 (kcal mol–1) | Kd1(µM) | ΔH1(kcal/mol) | ΔH2 (kcal mol–1) | Kd1(µM) | ΔH1(kcal/mol) | Kd(µM) | Kd1(µM) | ΔH1(kcal/mol) | ΔG (kcal mol–1) | |
| WT | 85 | –4.5 | –0.9 | –5.5 | 260 | –2.6 | -2 | –4.6 | 105 | –2.3 | –2.9 | –5.2 |
| 50 | –4.7 | –0.9 | –5.6 | 220 | –4.1 | –0.6 | –4.7 | 105 | -9 | 3.5 | –5.5 | |
| 30 | –2.9 | –2.9 | –5.8 | 180 | –4.7 | –0.3 | -5 | 115 | –7.6 | 2.1 | –5.5 | |
| 55±15 | –4.0±0.5 | –1.6±0.6 | –5.6±0.1 | 220±20 | –3.8±0.6 | –1.0±0.5 | –4.8±0.1 | 110±3 | 6±2 | –2.8±1.9 | –5.4±0.1 | |
| A47W | 220 | –4.4 | –0.2 | –4.6 | 130 | 0.6* | –5.6 | -5 | 165 | –4.8 | –0.2 | -5 |
| Did not fit | 140 | –3.3 | –1.7 | -5 | ||||||||
| 150±10 | –4.0±0.7 | –0.9±0.5 | –5.0±0.0 | |||||||||
| D56A | 100 | –6.1 | 0.6 | –5.5 | 3000 | –11.2 | 7.7 | –3.5 | 190 | –9.2 | 4.1 | –5.1 |
| 55 | –4.5 | –1.2 | –5.7 | 1500 | –6.2 | 2.3 | –3.9 | 130 | –6.9 | 1.5 | –5.4 | |
| 80±20 | –5.3±0.8 | –0.6±0.9 | –5.6±0.1 | 2250±750 | –8.7±2.5 | 5.0±0.5 | –3.7±0.1 | 160±30 | –8.0±1 | –2.8±1.3 | –5.2±0.1 | |
| M230A | Did not fit | 142 | –6.1 | 0.9 | –5.2 | |||||||
| 182 | –3.1 | –1.7 | –4.8 | |||||||||
| 160±20 | –4.6±1.5 | –0.4±0.3 | –5.0±0.2 | |||||||||
| G223W | No Binding | |||||||||||
| D296A | Did not fit | 122 | –3.5 | –1.8 | –6.8 | |||||||
| 120 | –3.7 | –1.5 | –5.2 | |||||||||
| 120±1 | –3.6±0.1 | –1.6±0.1 | –6.0±0.8 | |||||||||
| D369A | 100 | –1.6 | –3.5 | –5.1 | 5350 | 2.8* | –5.9 | –3.1 | 80 | –1.4 | –4.1 | –5.5 |
| 90 | –2.5 | –2.6 | –5.7 | 4850 | 5* | -8 | -3 | 60 | –2.1 | –3.8 | –5.9 | |
| 95±5 | –5.3±0.8 | –3.1±0.4 | –5.4±0.1 | 5100±250 | 3.9±1.0 * | –6.9±1.0 | –3.0±0.1 | 70±10 | –1.7±0.3 | –3.9±0.1 | –5.7±0.1 | |
| A47W-D296A | No Binding | |||||||||||
| A47W-D369A | No Binding | |||||||||||
| D56A-D296A | No Binding | |||||||||||
| D56A-D369A | No Binding | |||||||||||
| M230A-D296A | No Binding | |||||||||||
| M230A-D369A | No Binding | |||||||||||
*
Positive values of ΔH corresponding to an exothermic mode of Cd2+ binding to protein indicates inappropriate fit.