Human beta(2) glycoprotein I as an anticardiolipin cofactor determined using deleted mutants expressed by a baculovirus system
Human beta(2) glycoprotein I as an anticardiolipin cofactor determined using deleted mutants expressed by a baculovirus system
复制标题
DOI：
10.1182/blood.v87.8.3262.bloodjournal8783262
复制
发表时间：
1996-04-15
期刊：
影响因子：
20.3
通讯作者：
Koike, T
中科院分区：
文献类型：
--
作者：
Igarashi, M;Matsuura, E;Koike, T
β（2）-糖蛋白I（beta（2）-GPI）由5个重复的同源结构域组成。我们设计了一系列人β（2）-GPI突变基因，即3个缺失NH 2端结构域的突变基因和2个缺失COOH端结构域的突变基因。这些突变基因在重组杆状病毒感染的草地贪夜蛾昆虫细胞（Sf 9）中表达，并分泌到培养基中。纯化的突变体蛋白质的分子量，估计由十二烷基硫酸钠-聚丙烯酰胺凝胶电泳，是相当一致的大小从它们的核苷酸序列计算。β（2）-GPI与固相心磷脂（CL）的结合通过从其完整结构中删除第五个结构域（结构域V）而减弱。因此，β（2）-GPI的磷脂结合位点位于其结构域V上。来自NZW x BXSB（WB）F1小鼠或抗磷脂综合征患者的单克隆抗CL抗体（aCL）直接与结构域V缺失的突变蛋白（DI-IV）结合，不仅吸附在氧化的聚苯乙烯表面，而且吸附在普通聚苯乙烯表面。我们从这项研究中得出结论，aCL的表位通过与带负电荷的磷脂相互作用暴露在β（2）-GPI的构象变化结构上或暴露在突变蛋白DI-IV上。(C)1996年，美国血液学会。
beta(2)-Glycoprotein I (beta(2)-GPI) consists of five repeats of a homologous domain, We designed a series of human beta(2)-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 beta(2)-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 beta(2)-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 beta(2)-GPI by interacting with negatively charged phospholipid or on the mutant protein, DI-IV. (C) 1996 by The American Society of Hematology.