THE 5TH DOMAIN OF BETA(2)-GLYCOPROTEIN-I CONTAINS A PHOSPHOLIPID-BINDING SITE (CYS281-CYS288) AND A REGION RECOGNIZED BY ANTICARDIOLIPIN ANTIBODIES

We have identified a phospholipid binding site in the fifth domain of beta2-glycoprotein I (beta2-GPI). Using synthetic peptides spanning th e fifth domain of beta2-GPI, we have shown that the presence of the se quence Glu274-Cys288 caused a decrease in the binding of purified anti cardiolipin (aCL) antibodies in a modified cardiolipin (CL)-ELISA by i nhibiting the binding of beta2-GPI to CL. This peptide bound to and co uld be eluted from a CL affinity column in a manner similar to native beta2-GPI. Peptides corresponding to other regions of the fifth domain had no inhibitory effect. The inhibitory activity was restricted to t he sequence Cys281-Lys-Asn-Lys-Glu-Lys-Lys-Cys288. Peptides in which t he two flanking cysteine residues were deleted or substituted with ser ine residues possessed no inhibitory activity, indicating that the con formation of this highly positively charged sequence may be critical f or phospholipid binding. aCL antibodies purified from patients with au toimmune disease were shown to bind directly to wells coated with nati ve beta2-GPI but not to wells coated with a preparation of beta2-GPI c leaved between Lys317 and Thr318. The integrity of this sequence is th erefore critical for these antibodies to recognize beta2-GPI, and the putative epitope for aCL antibodies is most likely to be in this regio n.