title: Microcalorimetric investigation of the interactions in the ternary complex calmodulin-calcium-melittin.
abstract: Flow microcalorimetric titrations of calmodulin with melittin at 25 degrees C revealed that the formation of the high-affinity one-to-one complex in the presence of Ca2+ (Comte, M., Maulet, Y., and Cox, J. A. (1983) Biochem, J. 209, 269-272) is entirely entropy driven (delta H0 = 30.3 kJ X mol-1; delta S0 = 275 J X K-1 X mol-1). Neither the proton nor the Mg2+ concentrations have any significant effect on the strength of the complex. In the absence of Ca2+, a nonspecific calmodulin-(melittin)n complex is formed; the latter is predominantly entropy driven, accompanied by a significant uptake of protons and fully antagonized by Mg2+. Enthalpy titrations of metal-free calmodulin with Ca2+ in the presence of an equimolar amount of melittin were carried out at pH 7.0 in two buffers of different protonation enthalpy. The enthalpy and proton release profiles indicate that: protons, absorbed by the nonspecific calmodulin-melittin complex, are released upon binding of the first Ca2+; Ca2+ binding to the high-affinity configuration of the calmodulin-melittin complex displays an affinity constant greater than or equal to 10(7) M-1, i.e. 2 orders of magnitude higher than that of free calmodulin; the latter is even more entropy driven (delta H0 = 7.2 kJ X site-1; delta S0 = 158 J X K-1 X site-1) than binding to free calmodulin (delta H0 = 4.7 kJ X site-1; delta S0 = 112 J X K-1 X site-1), thus underlining the importance of hydrophobic forces in the free energy coupling involved in the ternary complex.
mesh_terms: D000818:Animals; D001514:Bee Venoms / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism*; D002151:Calorimetry; D002417:Cattle; D002850:Chromatography, Gel; D008274:Magnesium / Q000378:metabolism; D008555:Melitten / Q000378:metabolism*; D008956:Models, Chemical; D013816:Thermodynamics
chemical_list: D001514:Bee Venoms; D002147:Calmodulin; D008555:Melitten; D008274:Magnesium; D002118:Calcium