Interaction of annexins IV and VI with phosphatidylserine in the presence of Ca2+: monolayer and proteolytic study.

Annexins, Ca2+- and phospholipid-binding proteins are known to bind to artificial and biological membranes in a calcium-dependent manner. However, the precise mechanism of the annexin-membrane interactions still remains to be studied in detail. In this paper we describe the results of studies on the interactions of the annexin/Ca complexes with phospholipids, obtained by the Wilhelmy balance method of assessing the surface pressure of a phospholipid monolayer. We show that the annexin IV/Ca as well as annexin VI/Ca complexes significantly reduce the surface pressure of a phosphatidylserine monolayer, when its initial value is close to collapse pressure. The effect is highly specific for monolayers composed of phosphatidylserine and strongly sensitive to pH and ionic strength. The most pronounced changes have been observed at pH 7.0-7.5, at a protein/Ca molar ratio of 1:2 for annexin IV and 1:4 for annexin VI. In the presence of sodium chloride at concentrations exceeding 400mM this effect was almost completely abolished. The obtained results point to the mainly electrostatic character of the annexin/phosphatidylserine interactions. In addition, using large multilamellar lipid vesicles and serine proteases, we demonstrate that annexins, when bound in a ternary complex with phospholipids and calcium ions, are partially protected against proteolysis. Our observation that annexin molecules, complexed with calcium ions, are protected against proteolytic attack in the presence of PS liposomes does not have to be necessarily explained in terms of partial penetration of protein within the membrane bilayer.