Table 2.
Thermodynamic parameters of CPX binding to SNAREs measured by Isothermal Titration Calorimetry. Superclamp CPX (scCPX, residues 1–134 carrying superclamp mutation D27L, E34F, R37A, scCPX) or CPX48–134 were titrated into assembled SNARE complexes containing VAMP2 (residues 1–96) or VAMP-3xDA (residues 1–96 with mutations D64A, D65A, & D68A). The thermodynamic parameters, dissociation constant (Kd); enthalpy (ΔH); entropy (ΔS) and free energy (ΔG) were calculated by nonlinear least squares fit with a one-set-of-sites model from the binding isotherms shown in Fig. 3. Average and standard deviations of a minimum of three independent experiments are shown.
| Titrant | In sample cell | Stoichiometric Coefficient (N) | Kd (nM) | ΔH (kCal mol−1) | ΔS (Cal mol−1 K−1) | ΔG (kCal mol−1) |
|---|---|---|---|---|---|---|
| scCPX | SNARE complex with VAMP2-WT | 0.95 ± 0.01 | 83 ± 17 | −37.5 ± 0.7 | −88.6 ± 2.7 | −10.1 ± 0.2 |
| scCPX | SNARE complex with VAMP-3xDA | 0.99 ± 0.04 | 670 ± 90 | −15.0 ± 1.0 | −20.1 ± 3.6 | −8.8 ± 0.2 |
| CPX48–134 | SNARE complex with VAMP-3xDA | 0.98 ± 0.01 | 620 ± 110 | −14.0 ± 1.1 | −16.6 ± 3.8 | −8.8 ± 0.1 |