HUMAN BETA(2) GLYCOPROTEIN I AS AN ANTICARDIOLIPIN COFACTOR DETERMINED USING DELETED MUTANTS EXPRESSED BY A BACULOVIRUS SYSTEM

beta(2)-Glycoprotein I (beta(2)-GPI) consists of five repeats of a hom ologous domain, We designed a series of human beta(2)-GPI mutant genes , ie, three mutant genes lacking the domain(s) present in the NH2-term inal region and two of those present in the COOH-terminal region. Thes e mutant genes were expressed in Spodoptera frugiperda insect cells (S f9) infected with recombinant baculoviruses and the mutant proteins we re secreted into the culture medium. The molecular mass of the purifie d mutant proteins, estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, was fairly consistent with the size calculated fr om their nucleotide sequences. Binding of beta(2)-GPI to solid-phase c ardiolipin (CL) was diminished by the deletion of the fifth domain (do main V) from its complete structure. Thus, the phospholipid binding si te of beta(2)-GPI is located on its domain V. Monoclonal anti-CL antib odies (aCL) derived either from NZW x BXSB (WB) F1 mice or from patien ts with antiphospholipid syndrome bound directly to the domain V-delet ed mutant protein (DI-IV) absorbed not only on an oxygenated but also on a plain polystyrene surface. We conclude from this study that the e pitope for aCL is exposed on a comformationally changed structure of b eta(2)-GPI by interacting with negatively charged phospholipid or on t he mutant protein, DI-IV. (C) 1996 by The American Society of Hematolo gy.