Competition of annexin V and anticardiolipin antibodies for binding to phosphatidylserine containing membranes

Annexin V, an intracellular protein with a calcium-dependent high affinity for anionic phospholipid membranes, acts as an inhibitor of lipid-dependent reactions of the blood coagulation. Antiphospholipid antibodies found in t he plasma of patients with antiphospholipid syndrome generally do not inter act with phospholipid membranes directly, but recognize (plasma) proteins a ssociated with lipid membranes, mostly prothrombin or beta(2)-glycoprotein I (beta(2)GPI). Previously, it has been proposed that antiphospholipid anti bodies may cause thrombosis by displacing annexin V from procoagulant cell surfaces. We used ellipsometry to study the binding of annexin V and of com plexes of beta(2)GPI with patient-derived IgG antibodies to beta(2)GPI, com monly referred to as anticardiolipin antibodies (ACA), to phospholipid bila yers composed of phosphatidylcholine (PC) and 20% phosphatidylserine (PS). More specifically, we investigated the competition of these proteins for th e binding sites at these bilayers. We show that ACA-beta(2)GPI complexes, a dsorbed to PSPC bilayers, are displaced for more than 70% by annexin V and that annexin V binding is unaffected by the presence of ACA-beta(2)GPI comp lexes. Conversely, annexin V preadsorbed to these bilayers completely preve nts adsorption of ACA-beta(2)GPI complexes, and none of the preadsorbed ann exin V is displaced by ACA-beta(2)GPI complexes. Using ellipsometry, we als o studied the effect of ACA-beta(2)GPI complexes on the interaction of anne xin V with the membranes of ionophore-activated blood platelets as a more p hysiological relevant model of cell membranes. The experiments with blood p latelets confirm the high-affinity binding of annexin V to these membranes and unequivocally show that annexin V binding is unaffected by the presence of ACA-beta(2)GPI. In conclusion, our data unambiguously show that ACA-bet a(2)GPI complexes are unable to displace annexin V from procoagulant membra nes to any significant extent, whereas annexin V does displace the majority of preadsorbed ACA-beta(2)GPI complexes from these membranes.