Role of Divalency in the High-Affinity Binding of Anticardiolipin Antibody−β2-Glycoprotein I Complexes to Lipid Membranes

• Published in: Biochemistry (Easton) Vol. 35; no. 43; pp. 13833 - 13842
• Main Authors: Willems, George M, Janssen, Marie P, Pelsers, Maurice M. A. L, Comfurius, Paul, Galli, Monica, Zwaal, Robert F. A, Bevers, Edouard M
• Format: Journal Article
• Language: English
• Published: American Chemical Society 29.10.1996
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/bi960657q

β2-Glycoprotein I (β2GPI) is an essential cofactor for the binding to lipids of anticardiolipin antibodies (ACA), isolated from patients with anti-phospholipid syndrome. We used ellipsometry to study the binding of β2GPI and the β2GPI-mediated binding of ACA to planar membranes composed of phosphatidylcholine (PC) and 5−20 mol % phosphatidylserine (PS). No binding of β2GPI was observed to neutral (PC) membranes. Maximal binding of β2GPI was 3.2−3.6 pmol·cm-2. Affinity decreased strongly with decreasing PS content; increasing the NaCl and CaCl2 concentrations also led to a decrease in affinity. At physiologic conditions (10 mol % PS, 120 mM NaCl, and 3 mM CaCl2), a K d of 14 μM was observed. Binding constants were insensitive to the chemical composition of the negatively charged phospholipid headgroup. ACA (1.25−10 μg·mL-1) caused a 30−40-fold enhancement of β2GPI binding to PS/PC membranes (20 mol % PS), resulting in the binding of about 2 pmol·cm-2 divalent ACA−(β2GPI)2 complexes at 100 nM β2GPI. In the absence of β2GPI, binding of ACA was negligible. Ad- and desorption kinetics of ACA−β2GPI complexes indicate that the initial monovalent association of ACA to membrane-bound β2GPI is rapidly followed by formation of divalent ACA−(β2GPI)2 complexes. Experiments with monovalent Fab1 fragments of ACA showed no appreciable effect on the β2GPI binding to lipid, substantiating the notion that divalent interactions are essential for the high-affinity binding of ACA−β2GPI. The anticoagulant effect of ACA is rationalized by the observation that binding of ACA−β2GPI complexes to the PSPC membrane severely restricts the adsorption of blood coagulation factor Xa.