Isothermal titration calorimetric study of calcium association to lipid bilayers: influence of the vesicle preparation and composition.
The association of Ca2+ ions with phospholipid bilayers was investigated using isothermal titration calorimetry. The study reveals that the binding enthalpy of these cations to bilayers formed with 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG) depends strongly on the method of preparation of the unilamellar vesicles. Extruded vesicles lead to an exothermic association, whereas sonicated ones lead to an endothermic association. In the later case, the calorimetric signal is sensitive to the length of the sonication period. It is proposed that a reorganization of the lipid bilayers under stress, obtained with sonicated small unilamellar vesicles, contributes to the calorimetric signal upon the titration with Ca2+. The analysis of the titrations indicates that, as expected, the nature of the association of Ca2+ with negatively charged phospholipid bilayers is essentially of electrostatic nature. Using a Scatchard approach, it is found that bilayers become saturated in Ca2+ approximately when the electroneutrality of the bilayer interface is reached. Moreover, the affinity constant was reduced by the increase of the ionic strength of the aqueous buffer. It was found that the intrinsic binding constant of Ca2+ to membranes containing 30 and 50 mol% of POPG was about 11 mM-1, in a MES buffer containing 10 mM NaCl, at pH 5.6.
Arseneault
Marjolaine
M
Department of Chemistry, Université de Montréal, C.P. 6128, Succ. Centre Ville, Montréal, Que., H3C3J7 Canada.
Lafleur
Michel
M
D002118:Calcium / Q000737:chemistry* / Q000378:metabolism; D002151:Calorimetry / Q000379:methods; D006863:Hydrogen-Ion Concentration; D008051:Lipid Bilayers / Q000737:chemistry* / Q000378:metabolism; D008561:Membrane Fusion; D010713:Phosphatidylcholines / Q000737:chemistry* / Q000378:metabolism; D010715:Phosphatidylglycerols / Q000737:chemistry* / Q000378:metabolism; D013816:Thermodynamics; D013997:Time Factors; D017186:Titrimetry
D008051:Lipid Bilayers; D010713:Phosphatidylcholines; D010715:Phosphatidylglycerols; C060037:1-palmitoyl-2-oleoylglycero-3-phosphoglycerol; D002118:Calcium; C028694:1-palmitoyl-2-oleoylphosphatidylcholine