Table 1.
Determination of the dissociation constant values of lipid binding to the protein kinase Cα C2 domain using isothermal titration calorimetry measurement and comparison of the wild-type C2 domain to other forms that have been the subject of target-site mutations
| Protein | Lipida | Presence of Ca2+ | K d (μM) | Reference |
|---|---|---|---|---|
| WT-C2 | Soluble IP3 | + | 19.8 ± 1.7 | Guerrero-Valero et al. (2009) |
| WT-C2 | POPC/POPS (2:3 molar ratio) | + | 18.0 ± 1.2 | Torrecillas et al. (2004) |
| WT-C2 | POPC/POPS (2:3 molar ratio) | - | 430 ± 23 | Torrecillas et al. (2004) |
| WT-C2 | POPC/PIP2 (95:5 molar ratio) | + | 1.9 ± 0.4 | Sanchez-Bautista et al. (2006) |
| WT-C2 | POPC/PIP2 (95:5 molar ratio) | - | 48.3 ± 9.7 | Sanchez-Bautista et al. (2006) |
| C2 D246/248N | POPC/PIP2 (95:5 molar ratio) | + | 26.5 ± 5.0 | Sanchez-Bautista et al. (2006) |
| C2 K209/211A | POPC/PIP2 (95:5 molar ratio) | + | No binding detected | Sanchez-Bautista et al. (2006) |
| WT-C2 (two sets of sites) | POPC/POPS/PIP2 (70:25:5 molar ratio) | + | 1.6 ± 0.2 1.7 ± 0.4 |
Guerrero-Valero et al. (2007) |
| WT-C2 (one set of sites) | POPC/POPS/PIP2 (70:25:5 molar ratio) | + | 1.9 ± 0.1 | Guerrero-Valero et al. (2007) |
K d,Equilibrium dissociation constant; WT-C2, wild-type protein kinase C (PKC) domain;
When phospholipids were used, they were in the form of sonicated vesicles. When Ca2+ was present, it was added to reach saturating concentrations. In the last two cases, the origin for isothermal titration calorimetry (ITC) allowed fitting the results to two sets of binding sites model or to a one-site model
aIP3, Inositol 1,4,5-trisphosphate; POPC, phosphatidylcholine; PIP2, phosphatidylinositol-4,5-bisphosphate; POPS, phosphatidylserine