Studies on the mechanism of membrane fusion: evidence for an intermembrane Ca2+-phospholipid complex, synergism with Mg2+, and inhibition by spectrin.
The interaction of Ca2+ and Mg2+ with phosphatidylserine (PS) vesicles in 0.1 M NaCl aqueous solution was studied by equilibrium dialysis binding, X-ray diffraction, batch microcalorimetry, kinetics of cation-induced vesicle aggregation, release of vesicle contents, and fusion. Addition of either cation causes aggregation of PS vesicles and produces complexes with similar stoichiometry (1:2 cation/PS) at saturating concentrations, although the details of the interactions and the resulting complexes are quite different. Addition of Ca2+ to PS vesicles at T greater than or equal to 25 degrees C induces the formation of an "anhydrous" complex of closely apposed membranes with highly ordered crystalline acyl chains and a very high transition temperature (Tc greater than 100 degrees C). The formation of this complex is accompanied by a release of heat (5.5 kcal/mol), rapid release of vesicle contents, and fusion of the vesicles into larger membranous structures. By contrast, addition of Mg2+ produces a complex with PS which is much more hydrated, has no crystallization of the acyl chains at T greater than or equal to 20 degrees C, and has comparatively little fusion. Studies with both Ca2+ and Mg2+ added simultaneously indicate that there is a synergistic effect between the two cations, which results in an enhancement of the ability of Ca2+ to form its specific complex with PS at lower concentrations. The presence of the erythrocyte protein "spectrin" inhibits this synergism and interferes with the formation of the specific PS/Ca complex. It also inhibits the fusion of PS vesicles. It is proposed that the unique PS/Ca complex, which involves close apposition of vesicle membranes, is an intermembrane "trans" complex. We further propose that such a complex is a key step for the resultant phase transition and fusion of PS vesicles. By contrast, the PS/Mg complex is proposed to be a "cis" complex with respect to each membrane. The results are discussed in terms of the mechanism of membrane fusion.
D002118:Calcium; D002151:Calorimetry; D055598:Chemical Phenomena; D002621:Chemistry; D007700:Kinetics; D008081:Liposomes; D008274:Magnesium; D008565:Membrane Proteins; D010539:Permeability; D010718:Phosphatidylserines; D013049:Spectrin; D013499:Surface Properties; D013816:Thermodynamics; D014961:X-Ray Diffraction
D008081:Liposomes; D008565:Membrane Proteins; D010718:Phosphatidylserines; D013049:Spectrin; D008274:Magnesium; D002118:Calcium