Dimers of beta 2-glycoprotein I mimic the in vitro effects of beta 2-glycoprotein I-anti-beta 2-glycoprotein I antibody complexes.
Anti-beta(2)-glycoprotein I antibodies are thought to cause lupus anticoagulant activity by forming bivalent complexes with beta(2)-glycoprotein I (beta(2)GPI). To test this hypothesis, chimeric fusion proteins were constructed of the dimerization domain (apple 4) of factor XI and beta(2)GPI. Both a covalent (apple 4-beta(2)GPI) and a noncovalent (apple 4-C321S-beta(2)GPI) chimer were constructed. As controls, apple 2-beta(2)GPI and apple 4-C321S-beta(2)GPI-W316S, in which beta(2)GPI-W316S is not able to bind to phospholipids, were made. In a phospholipid binding assay, apple 4-beta(2)GPI and apple 4-C321S-beta(2)GPI were able to bind to phospholipids with an affinity 35 times higher than that of plasma-derived beta(2)GPI and apple 2-beta(2)GPI. Apple 4-C321S-beta(2)GPI-W316S did not bind at all. Only apple 4-beta(2)GPI and apple 4-C321S-beta(2)GPI were able to bind to adhered platelets as shown by immunofluorescence. Using the prothrombin time, which was the most responsive coagulation assay, the clotting time was approximately doubled when 200 microg/ml apple 4-beta(2)GPI or apple 4-C321S-beta(2)GPI was added. Addition of 200 microg/ml plasma-derived beta(2)GPI, apple 2-beta(2)GPI, or apple 4-C321S-beta(2)GPI-W316S did not affect clotting time. Clotting time could be corrected with the addition of extra phospholipids, which is indicative for lupus anticoagulant activity. An additional increase in clotting times for apple 4-beta(2)GPI or apple 4-C321S-beta(2)GPI was achieved by the addition of monoclonal antibodies against beta(2)GPI. In conclusion, dimerization of beta(2)GPI explains the in vitro observed effects of beta(2)GPI-anti-beta(2)GPI antibody complexes.
Lutters
B C
BC
Department of Haematology, University Medical Center, 3508 GA Utrecht, Utrecht University, 3508 TB Utrecht, The Netherlands. b.c.h.lutters@lab.azu.nl
Meijers
J C
JC
Derksen
R H
RH
Arnout
J
J
de Groot
P G
PG
D000818:Animals; D000936:Antigen-Antibody Complex / Q000378:metabolism*; D001780:Blood Coagulation Tests; D001792:Blood Platelets / Q000378:metabolism; D002478:Cells, Cultured; D002850:Chromatography, Gel; D006224:Cricetinae; D019281:Dimerization; D006023:Glycoproteins / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D016682:Lupus Coagulation Inhibitor / Q000378:metabolism; D010743:Phospholipids / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000378:metabolism; D019685:Rosales; D053482:beta 2-Glycoprotein I
D000936:Antigen-Antibody Complex; D006023:Glycoproteins; D016682:Lupus Coagulation Inhibitor; D010743:Phospholipids; D011993:Recombinant Fusion Proteins; D053482:beta 2-Glycoprotein I