Studies on the mechanism of membrane fusion: role of phosphate in promoting calcium ion induced fusion of phospholipid vesicles

Abstract

The role of phosphate in enhancing the Ca-induced fusion of phosphatidylserine (PS) vesicles was examined by using the new fluorescent (terbium/dipicolinic acid) assay described by Wilschut et al. (1980). In the presence of physiological levels of phosphate the Ca concentration required for fusion of PS vesicles was lowered significantly (3- to 4-fold), and the rate of vesicle fusion was increased dramatically (up to 1000-fold). The fusion of PS vesicles by Ca and phosphate is specific and critically dependent on temperature, pH, ion concentrations and the composition of the Ca phosphate crystalline phase present during the incubation. A significant enhancement in vesicle fusion apparently occurs only when Ca phosphate precipitation is initiated in the presence of PS vesicles, suggesting that crystal nucleation on the vesicle surface is a prerequisite for fusion. Ca and phosphate promote phospholipid phase separations and vesicle fusion under conditions (e.g., mixtures of PS and phosphatidylcholine) in which Ca alone is ineffective, indicating that formation of PS Ca phosphate complexes may facilitate the molecular segregation of PS into distinct domains. These experiments underline the important role that phosphate may play in Ca-mediated fusion phenomena in biological membranes.