Human β2-glycoprotein I as an anticardiolipin cofactor determined using deleted mutants expressed by a baculovirus system

Makoto Igarashi, Eiji Matsuura, Yoshiko Igarashi, Hisato Nagae, Kenji Ichikawa, Douglas A. Triplett, Takao Koike

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Abstract

β2-Glycoprotein I (β2-GPI) consists of five repeats of a homologous domain. We designed a series of human β2-GPI mutant genes, ie, three mutant genes lacking the domain(s) present in the NH2-terminal region and two of those present in the COOH-terminal region. These mutant genes were expressed in Spodoptera frugiperda insect cells (Sf9) infected with recombinant baculoviruses and the mutant proteins were secreted into the culture medium. The molecular mass of the purified mutant proteins, estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, was fairly consistent with the size calculated from their nucleotide sequences. Binding of β2- GPI to solid-phase cardiolipin (CL) was diminished by the deletion of the fifth domain (domain V) from its complete structure. Thus, the phospholipid binding site of β2-GPI is located on its domain V. Monoclonal anti-CL antibodies (aCL) derived either from NZW x BXSB (WB) F1 mice or from patients with antiphospholipid syndrome bound directly to the domain V-deleted mutant protein (DI-IV) absorbed not only on an oxygenated but also on a plain polystyrene surface. We conclude from this study that the epitope for aCL is exposed on a comformationally changed structure of β2-GPI by interacting with negatively charged phospholipid or on the mutant protein, DMV.

Original languageEnglish
Pages (from-to)3262-3270
Number of pages9
JournalBlood
Volume87
Issue number8
Publication statusPublished - Apr 15 1996
Externally publishedYes

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ASJC Scopus subject areas

  • Hematology

Cite this

Igarashi, M., Matsuura, E., Igarashi, Y., Nagae, H., Ichikawa, K., Triplett, D. A., & Koike, T. (1996). Human β2-glycoprotein I as an anticardiolipin cofactor determined using deleted mutants expressed by a baculovirus system. Blood, 87(8), 3262-3270.