The exchanged EF-hands in calmodulin and troponin C chimeras impair the Ca2+-induced hydrophobicity and alter the interaction with Orai1: a spectroscopic, thermodynamic and kinetic study

Table 1 ANS binding to CaM and chimeras by fluorescence and ITC

	ANS binding in the absence of salt		ANS binding in the presence of salt		ANS binding in the presence of salt and Orai -CMBD
	30 mM Hepes, pH 7.5, 2 mM Ca ²⁺		30 mM Hepes, pH 7.5, 0.1 M NaCl, 2 mM Ca ²⁺		30 mM Hepes, pH 7.5, 0.1 M NaCl, 2 mM Ca ²⁺
Protein	ANS enhancement (fold) ^a	K _app (M ⁻¹ )	ANS fold enhancement (fold) ^a	K _app (M ⁻¹ )	ANS fold enhancement (fold) ^a	K _app (M ⁻¹ )
CaM	3.01 ± 0.12	2.10 ± 0.11 × 10³	3.49 ± 0.07	3.69 ± 0.08 × 10³	3.87 ± 0.10	3.43 ± 0.65 × 10³
CaM(1TnC)	2.72 ± 0.10	9.86 ± 1.24 × 10²	3.33 ± 0.06	1.13 ± 0.04 × 10³	3.31 ± 0.05	1.20 ± 0.08 × 10³
CaM(2TnC)	2.55 ± 0.05	1.01 ± 0.07 × 10³	3.01 ± 0.14	1.10 ± 0.01 × 10³	3.07 ± 0.04	9.70 ± 0.31 × 10²
CaM(3TnC)	1.56 ± 0.04	6.70 ± 0.51 × 10²	1.74 ± 0.08	8.56 ± 0.21 × 10²	2.24 ± 0.11	1.07 ± 0.10 × 10³
CaM(4TnC)	2.37 ± 0.22	1.80 ± 0.05 × 10³	2.90 ± 0.19	2.02 ± 0.03 × 10³	3.62 ± 0.15	2.67 ± 0.24 × 10²

^aThe ANS intensity enhancements were calculated from spectra integration and compared to the corresponding apo proteins in identical experimental conditions.

ISSN: 1471-2091